Beta-hairpin peptidomimetic with elastase inhibitory activity and aerosol dosage forms thereof

ABSTRACT

The present invention relates to pharmaceutical aerosols comprising a β-hairpin peptidomimetic of formula cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-DPro-Pro-), or a pharmaceutically acceptable salt thereof, having inhibitory activity against human neutrophil elastase. It further relates to solid or liquid pharmaceutical compositions and kits for preparing and administering such aerosols. The invention can be used for the prevention, management or treatment of pulmonary diseases, such as alpha-1 antitrypsin deficiency (AATD), cystic fibrosis (CF), non-cystic fibrosis bronchiactasis (NCFB), or chronic obstructive pulmonary disease (COPD), or infections, or diseases, or conditions of the lungs, being mediated by or resulting from human neutrophil elastase activity. Thus, the invention further relates to a pharmaceutical composition or a pharmaceutical aerosol comprising the active compound cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-DPro-Pro-), or any pharmaceutically acceptable salt thereof, for use in a method for the prevention, management or treatment of diseases or conditions of the lungs being mediated by or resulting from human neutrophil elastase activity in a subject.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a Continuation application of co-pending applicationSer. No. 16/305,714, filed on Nov. 29, 2018, which is the National Phaseunder 35 U.S.C. § 371 of International Application No.PCT/EP2017/025157, filed on May 31, 2017, which claims the benefit under35 U.S.C. § 119(a) to patent application Ser. No. 16/020,210.7, filed inEurope on May 31, 2016, all of which are hereby expressly incorporatedby reference into the present application.

FIELD OF THE INVENTION

The invention relates to pharmaceutical aerosols comprising a β-hairpinpeptidomimetic of formulacyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), or apharmaceutically acceptable salt thereof, being specifically disclosedin WO2006/087001 A1 and having inhibitory activity against humanneutrophil elastase. It further relates to solid or liquidpharmaceutical compositions and kits for preparing and administeringsuch aerosols. The invention can be used for the prevention, managementor treatment of diseases or conditions of the lungs being mediated by orresulting from human neutrophil elastase activity, e.g. pulmonarydiseases, such as alpha-1 antitrypsin deficiency (AATD), cystic fibrosis(CF), non-cystic fibrosis bronchiactasis (NCFB), or chronic obstructivepulmonary disease (COPD), or infections of the lungs causing diseases orconditions of the lungs, being mediated by human neutrophil elastaseactivity. Thus, the invention further relates to a pharmaceuticalcomposition or a pharmaceutical aerosol comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, for use in a method forthe prevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject.

BACKGROUND OF THE INVENTION

Human neutrophil elastase (human NE), a member of the serine proteasefamily, constitutes an important therapeutic target. Besides cathepsin Gand proteinase 3 it is intimately involved in the modulation ofactivities of cytokines and their receptors. Particularly at sites ofinflammation, high concentration of human NE is released frominfiltrating polymorphonuclear cells in close temporal correlation toelevated levels of inflammatory cytokines, strongly indicating that thisprotease is involved in the control of cytokine bioactivity andavailability (U. Bank, S. Ansorge, J. Leukoc. Biol. 2001, 69, 177-90).Human NE is known to be mainly responsible for extracellular proteolysisand contributes to tissue damage by catalyzing the hydrolysis of a widevariety of matrix macromolecules, plasma proteins, inflammatorymediators, and cell surface receptors with important local and systemicconsequences (C. A. Oven; E. J. Campbell; J. Leukoc. Biol. 1999, 65,137-150). Thus, inhibitors of human NE are valuable novel drugcandidates for infectious inflammatory diseases, including lung diseaseslike chronic obstructive pulmonary disease (COPD), acute respiratorydistress syndrome (ARDS), cystic fibrosis (CF) and ischemic-reperfusioninjury, (H. Ohbayashi, Expert Opin. Investig. Drugs 2002, 11, 965-980;B. Korkmaz, T. Moreau, F. Gauthier, Biochimie 2008, 90, 227). They canmeet a significant need for new therapies effectively preventing ortreating and/or mitigating these diseases or conditions.

Routes of administration can be classified whether the effect of thedrug is local (topical administration) or systemic (enteral orparenteral administration). The delivery of pharmaceuticals to thebronchi and lungs (pulmonary drug delivery) has been used for the localtreatment of diseases and conditions of the respiratory system. However,the feasibility of inhalation as an alternate route of administrationfor treatment of systemic diseases utilising the large surface of thelungs for absorption has been demonstrated as well (J. S. Patton, P. R.Byron, Nat. Rev. Drug Discov. 2007, 6, 67-74; M. Hohenegger, Curr.Pharm. Des. 2010, 16, 2484-2492).

In particular, drug substances can be delivered to the respiratorysystem as aerosolized dry powders or aerosolized liquids, the latterbeing either solutions or dispersions, such as drug substancesuspensions. Various devices have been developed to convert a solid orliquid composition into an aerosol to enable inhalation. For theconversion of aqueous-based drug substance solutions or suspensions intoinhalable aerosols nebulizers are normally used. They are particularlyuseful for diseases that require high pulmonary doses, e.g. CF, andpatients, e.g. children, who are unable to coordinate or achieve flowrates necessary for use of other inhalation devices (M. Knoch, M.Keller, Expert Opin. Drug Del. 2005, 2, 377-390).

Inhalation of drug substances may be favourable for the prophylaxisand/or treatment of respiratory tract infections and/or diseasesassociated with neutrophil elastase activity. There is evidence thatairway infiltration by neutrophils will lead to an increase inneutrophil elastase within the extracellular compartment and therebyinducing deleterious effects. In a mouse model of neutropil allergenchallenge it could be demonstrated that the inhibition of neutrophilelastase by sivelelstat attenuates airway hyperresponsiveness andinflammation (H. Koga, N. Miyahara et al., Respir. Res. 2013, 14:8). Ina different mouse model delivery of unglycosylated rhalpha1PI, arecombinant form of the elastase inhibitor alpha-1-proteinase inhibitor(alpha1PI), to the airway surface of CD-1 mice by nasal instillation wasfound to be highly protective against elastase-mediated injury, evensignificantly more protective than glycosylated blood-derived alpha1PI.These results suggest that aerosol delivery of rhalpha1PI could be aneffective strategy for controlling elastase-dependent pathophysiologyassociated with cystic fibrosis lung disease. Recently, R. Siekmeiersummarized the results of so far completed clinical studies aiming atlung deposition of inhaled alpha1PI of patients with hereditaryalpha1PI-deficiency or cystic fibrosis (Eur. J. Med. Res. 2010, 15[Suppl. II], 164). For both therapeutic areas most of the studiesindicate that the administration of alpha1PI by inhalation may serve asa potent therapy to neutralize the excess of inflammatory proteins andneutrophil elastase. Whereas the latter effects do not automaticallylead to improvements on patient's lung function, a clear reduction ofairway inflammation after alpha1PI treatment may precede pulmonarystructural changes (M. Griese, P. Latzin et al., Eur. Respir. J. 2007,29, 240).

Generally, the benefit of drug substance delivery via inhalation is thatit can afford delivery of sufficient therapeutic dosages of the drugdirectly to the primary site of action, e.g. in case of respiratorydiseases, while minimizing the risks of systemic toxicity. Additionally,suboptimal pharmacokinetics and/or pharmacodynamics associated withsystemic drug exposure may be avoided. Furthermore, inhalation (at home)is a more convenient mode of administration than intravenous injection(medical ward).

Still, the effectiveness of a therapy based on inhaled drug substancedelivery depends mainly on the drug which is selected, a pharmaceuticalcomposition thereof suitable for inhalation and the device that isemployed. Thus, there is a strong need for further pharmaceuticalcompositions, aerosols and therapeutic kits which are suitable for theprevention, management or treatment of diseases or conditions beingmediated by or resulting from human neutrophil elastase activityimproving the outcome of presently known therapies and/or overcoming thedisadvantages of presently known therapies.

SUMMARY OF THE INVENTION

The invention provides a pharmaceutical aerosol for pulmonaryadministration comprising a dispersed liquid phase and a continuous gasphase. The dispersed liquid phase comprises aqueous droplets comprisingthe active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof. The droplets of thedispersed phase have a mass median diameter from about 1.5 μm to about 5μm with a droplet size distribution having a geometrical standarddeviation from about 1.2 to about 1.7. Further provided by the inventionis the pharmaceutical aerosol for pulmonary administration comprising adispersed liquid phase and a continuous gas phase, wherein the dispersedliquid phase comprises aqueous droplets comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, for use in a method forthe prevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject.

In another aspect, the invention provides liquid and solidpharmaceutical compositions from which the above aerosol can beprepared. The liquid composition comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in a concentration withina range from about 4 mg/mL to about 100 mg/mL, preferably, within arange from about 17 mg/mL to about 95 mg/mL, or about 35 mg/mL to about95 mg/mL, respectively, and more preferably, within a range from about70 mg/mL to about 95 mg/mL.

In still another aspect, the invention provides a kit comprising anebulizer and a liquid or solid composition, wherein the nebulizer isadapted to aerosolize the liquid composition into an aerosol, asdescribed above. Further provided by the invention is a kit comprising anebulizer and a liquid or solid composition, wherein the nebulizer isadapted to aerosolize the liquid composition into an aerosol, asdescribed above, for use in a method for the prevention, management ortreatment of diseases or conditions of the lungs being mediated by orresulting from human neutrophil elastase activity in a subject.

In still another aspect, the invention further discloses a method ofpreparing and delivering an aerosol for pulmonary administration. Themethod comprises the step of providing a liquid pharmaceuticalcomposition comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in a concentration withina range from about 4 mg/mL to about 100 mg/mL, preferably, within arange from about 17 mg/mL to about 95 mg/mL, or about 35 mg/mL to about95 mg/mL, respectively, and more preferably, within a range from about70 mg/mL to about 95 mg/mL, or providing a solid pharmaceuticalcomposition for preparing the liquid composition, wherein thecomposition comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and wherein the solidcomposition is dissolvable or dispersible in an aqueous liquid solvent,and wherein the liquid composition comprises a concentration within arange from about 4 mg/mL to about 100 mg/mL, preferably, within a rangefrom about 17 mg/mL to about 95 mg/mL, or about 35 mg/mL to about 95mg/mL, respectively, and more preferably, within a range from about 70mg/mL to about 95 mg/mL, of the active compound, or any pharmaceuticallyacceptable salt thereof, and the step of providing a nebulizer capableof aerosolizing said liquid pharmaceutical composition at a meandelivery rate of at least about 0.8 mg of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, per minute, the nebulizerbeing further adapted to emit an aerosol comprising a dispersed liquidphase having a mass median diameter from about 1.5 μm to about 5 μm, andhaving a droplet size distribution having a geometrical standarddeviation from about 1.2 to about 1.7. In a subsequent step thenebulizer is operated to aerosolize the liquid pharmaceuticalcomposition which finally can be inhaled by mammals, more preferably, byhuman subjects.

In still another aspect, the invention provides a pharmaceuticalcomposition comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and optionally one or morepharmaceutically acceptable diluents, excipients or carriers, for use ina method for the prevention, management or treatment of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject.

In still another aspect, the invention provides a kit comprising theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and a package insertwherein the package insert comprises instructions for treating a subjectfor diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity using the active compound.

BRIEF DESCRIPTION OF THE FIGURES

FIGS. 1 and 2 show the effects ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation in a LPS/fMLP model of neutrophil activationin the rat. BAL supernatants from the tracheae of dead rats wereanalyzed for neutrophil elastase activity. Data are displayed versus thevehicle control. Data are depicted both as means (FIG. 1 ) and asindividual data points together with their corresponding mean and s.e.m.values (FIG. 2 ).

FIG. 3 shows the mean plasma concentration-time curves of singleascending doses (80 mg, 160 mg and 320 mg per cohort) ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation to patients with cystic fibrosis. Thesecurves show typical profiles similar to profiles of curves resultingfrom comparable doses of active compound inhaled by healthy subjects.

FIG. 4 shows the mean sputum concentrations ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation to patients with cystic fibrosis as singleascending doses (80 mg, 160 mg and 320 mg per cohort). Data aredisplayed versus time. Comparison of concentrations of active compoundin sputum (FIG. 4 ) and plasma (FIG. 3 ) shows that concentrations ofactive compound in sputum are approximately 10³ fold higher than inplasma.

FIG. 5 shows the effects on active neutrophil elastase in sputum ofpatients with cystic fibrosis induced bycyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation to patients with cystic fibrosis as singleascending doses (80 mg, 160 mg and 320 mg per cohort). The data aredisplayed versus time and depicted as means. Additionally, data ofplacebo are depicted as well.

Strong inhibition (>90%) of active elastase in sputum over several hoursafter single dose administration of active compound in all dose groupscan be derived.

DETAILED DESCRIPTION OF THE INVENTION

In a first aspect, the invention provides a pharmaceutical aerosol forpulmonary administration comprising a dispersed liquid phase and acontinuous gas phase. The dispersed liquid phase comprises aqueousdroplets comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof. The droplets of thedispersed phase have a mass median diameter from about 1.5 μm to about 5μm with a droplet size distribution having a geometrical standarddeviation from about 1.2 to about 1.7.

The aerosol of the invention is for pulmonary delivery, which ispreferable achieved via oral inhalation of the aerosol. As used hereinin the description and the claims, pulmonary delivery means aerosoldelivery to any part or feature of the lungs including the so-calleddeep lungs, the peripheral lungs, the alveoli, the bronchi and thebronchioli.

Conditions of the pulmonary target regions in which the prevention,management or treatment of mammals, more preferably, of human subjects,using the aerosol of the invention is potentially useful includediseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity, e.g. in particular, pulmonarydiseases, such as alpha-1 antitrypsin deficiency (AATD), cystic fibrosis(CF), non-cystic fibrisis bronchiactasis (NCFB), or chronic obstructivepulmonary disease (COPD), or infections of the lungs causing diseases orconditions of the lungs, being mediated by human neutrophil elastaseactivity.

As used herein in the description and the claims, an aerosol is adispersion of a solid and/or liquid phase in a gas phase. The dispersedphase, also termed discontinuous phase, comprises multiple solid and/orliquid particles. Both basic physical types of aerosols, i.e. solid andliquid dispersions in a gas phase, may be used as pharmaceuticalaerosols.

According to the present invention, the aerosol comprises a dispersedliquid phase and a continuous gas phase. Such aerosols are sometimesreferred to as “liquid aerosols” or aerosolized liquids. It should benoted that the requirement of a dispersed liquid phase does not excludethe presence of a solid phase. In particular, the dispersed liquid phasemay itself represent a dispersion, such as a suspension of solidparticles in a liquid.

The continuous gas phase is to be selected from any gas or mixture ofgases which is pharmaceutically acceptable. For example, air orcompressed air as gas phase is most common in inhalation therapy usingnebulizers as aerosol generators. Alternatively, other gases and gasmixtures, such as air enriched with oxygen, or mixtures of nitrogen andoxygen may be used. The use of air as continuous gas phase is mostpreferred.

The active compound iscyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, having inhibitory activityagainst human neutrophil elastase. Structurally, the active compound isa homodetic, cyclic tridecapeptide, wherein OctG is (S)-2-aminodecanoicacid and ^(D)Pro is D-proline. The abbreviations (3-letter code) for theremaining amino acid residues are as generally recognized. All aminoacid residues are in L-configuration except one D-proline residue.

As used herein in the description and the claims the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) shouldbe understood as to include the respective solvates.

Solvates as well as salts are categories of forms in which the activecompoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) may beused as an active ingredient in a pharmaceutical composition.

Salts are neutral compounds composed of ions, i.e. cations and anions.If the active compound can act like an acid, potentially useful saltsmay be formed with inorganic cations, such as sodium, potassium,calcium, magnesium and/or ammonium, or with organic cations, such asthose derived from arginine, lysine, glycine, and/or ethylenediamine. Ifthe active compound (or parts thereof) can act like a base, as forexample the residue of Lys being one of the amino acid residues ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), thenpotentially useful salts may be formed with inorganic anions, such aschloride, bromide, iodide, phosphate (mono- or dibasic), sulfate,nitrate, acetate, trifluoroacetate, propionate, butyrate, maleate,fumarate, methanesulfonate, ethanesulfonate, 2-hydroxy-ethylsulfonate,n-propylsulfonate, isopropylsulfonate, lactate, malate, and/or citrate.

The term pharmaceutically acceptable salt or pharmaceutical salt is usedto refer to an ionisable drug or active compound that has been combinedwith a counter ion to form a neutral complex. Converting a drug oractive compound into a salt through this process can, for example,increase its chemical stability, render the complex easier to administerand/or allow manipulation of the agent's pharmacokinetic profile.

In a preferred embodiment of the invention, the counter ion of theactive compound is acetate.

The aerosol is further characterized in that the droplets of thedispersed liquid phase have a mass median diameter from about 1.5 μm toabout 5 μm with a droplet size distribution having a geometricalstandard deviation from about 1.2 to about 1.7. The mass median diameter(MMD), as used herein in the description and the claims, is the massmedian diameter of the dispersed liquid phase as measured by laserdiffraction. Various appropriate analytical apparatuses to determine theMMD are known and commercially available, such as the MalvernMasterSizer X or Malvern SprayTec. The geometric distribution includingthe geometric standard deviation (GSD) of the aerosolized liquidparticles or droplets may be determined simultaneously with the MMD. TheGSD describes how spread out is a set of numbers the preferred averageof which is the geometric mean.

In a preferred embodiment, the aerosol is for pulmonary delivery and thedispersed liquid phase of such an aerosol has a MMD in the range fromabout 2.0 μm to about 4.5 μm and a GSD in the range from about 1.2 toabout 1.7. More preferably, the aerosol of the invention has a MMD inthe range from about 2.5 μm to about 3.5 μm and a GSD in the range fromabout 1.4 to about 1.6. Each of these sets of combinations isparticularly useful to achieve a high local concentration of the activecompound in the lungs, including the bronchi and bronchioli, relative tothe amount of active compound which is aerosolized.

In another preferred embodiment, the aerosol is emitted from an aerosolgenerator at a rate of at least about 0.1 mL/min. In another embodiment,the (total) output rate being the rate at which the aerosol is emittedfrom the aerosol generator is at least about 0.150 mL/min or at leastabout 150 mg/min for those liquid aerosols the densities of whichare—for practical purposes—close to 1 g/mL, i.e. within the range fromabout 0.95 g/mL to about 1.05 g/mL. In further embodiments, the outputrate is within the range from about 200 mg/min to about 700 mg/min, orfrom about 250 mg/min to about 650 mg/min, respectively.

In another preferred embodiment, the aerosol is emitted from an aerosolgenerator at a mean delivery rate of at least about 0.8 mg of the activecompoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, per minute. The (mean)delivery rate of a drug or active compound is one of two discretemetrics or parameters being defined and measured according to e.g. Ph.Eur. (Pharmacopeia Europaea) 2.9.44 and/or USP (United StatesPharmacopeia) 1601 to determine the amount of drug or active compound apatient might be expected to receive during a treatment period. Infurther embodiments, the mean delivery rate of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, is within the range fromabout 3 mg to about 25 mg per minute or within the range from about 5 mgto about 18 mg per minute, respectively.

Appropriate aerosol generators, in particular nebulizers, which aresuitable for generating the aerosol(s) described herein in thedescription and the claims are discussed in more detail herein-below.

In another aspect, the present invention is directed to a liquidpharmaceutical composition for preparing the aerosol as described abovecomprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in a concentration withina range from about 4 mg/mL to about 100 mg/mL.

As defined herein in the description and the claims, a liquidpharmaceutical composition is a liquid material which comprises at leastone active compound and at least one pharmaceutically acceptable,pharmacologically substantially inert excipient. It should be noted thatthe term “liquid composition” does not necessarily mean that no solidmaterial is present. For example, a liquid suspension representing adispersion of solid particles in a continuous liquid phase is alsoembraced in the above term.

Preferably, the liquid composition from which the aerosol is prepared isan aqueous composition; consequently, water is the predominant liquidconstituent of such composition. Solvents and co-solvents other thanwater should be avoided. In another embodiment, the compositioncomprises at least about 80 wt.-% of water. In yet another embodiment,at least about 90 wt.-% of the liquid constituents of the composition iswater.

If the incorporation of a non-aqueous solvent, such as ethanol,glycerol, propylene glycol or polyethylene glycol, cannot be avoided,the excipient should be selected carefully and in consideration of itsphysiological acceptability and the therapeutic use of the composition.According to a preferred embodiment, the composition is substantiallyfree of non-aqueous solvents.

The concentration of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in the liquid compositionis within a range from about 4 mg/mL to about 100 mg/mL. Preferably, theconcentration of the above active compound, or any pharmaceuticallyacceptable salt thereof, is within the range from about 17 mg/mL toabout 95 mg/mL, or about 35 mg/mL to about 95 mg/mL, respectively, or,more preferred, about 70 mg/mL to about 95 mg/mL. A high concentrationof an active compound in a composition suitable to be aerosolizedprovides for patient convenience and compliance: The higher suchconcentration the smaller the total volume of liquid compositioncomprising the effective dose of the active compound to be inhaled andthe shorter the total time being necessary for inhalation of such aneffective dose.

The dynamic viscosity of the liquid composition has an influence on theefficiency of nebulization and on the particle size distribution of theaerosol formed by nebulization. The dynamic viscosity should preferablybe adjusted to a range from about 0.8 mPas to about 1.7 mPas. In otherembodiments, the dynamic viscosity is in the range from about 1.0 mPasto about 1.7 mPas, or in the range from about 1.2 mPas to about 1.6mPas, respectively.

In order to obtain an aerosol which is highly suitable for pulmonaryadministration, the surface tension of the liquid composition of theinvention should preferably be adjusted to a range from about 25 mN/m toabout 80 mN/m, more preferably, to a range from about 30 mN/m to about70 mN/m, or even more preferably, to a range from about 45 mN/m to about55 mN/m.

In general, the quality of an aerosol and the efficiency of thenebulisation could be adversely affected in the lower parts of the abovepresented embodiments; however, the results of the studies describedbelow indicate that there are no significant changes in the performancesof the liquid compositions of the invention in the above respect.

It is well known in the art that addition of a surfactant to an aqueousliquid composition may result in a surface tension being reduced fairlymarkedly below that of water or physiological buffer solution.Therefore, a compromise has to be found in each case depending on theintended application.

In order to be well-tolerated an aerosol should—as far as possible—havea physiological tonicity or osmolality. Thus, it may be desirable toincorporate an osmotically active excipient to control the osmolality ofthe aerosol. Such an excipient, or excipients, if e.g. a combination ofsubstances is used, should be selected to ideally reach an osmolality ofthe aerosol which does not deviate too much from that of physiologicalfluids, i.e. from about 150 mOsmol/kg. However, a compromise has againto be found between the physical-chemical and/or pharmaceutical needs onone hand and the physiological requirements on the other hand. Ingeneral, an osmolality up to about 800 mOsmol/kg may be acceptable. Inparticular, an osmolality in the range from about 200 mOsmol/kg to about600 mOsmol/kg is preferred. In further embodiments, the osmolality is inthe range from about 250 mOsmol/kg to about 500 mOsmol/kg or in therange from about 300 mOsmol/kg to about 450 mOsmol/kg, respectively.

One approach to improve the effectiveness and/or efficacy of thecomposition may be to enhance the local retention time of thecomposition after deposition of the aerosol in the target regions. Forexample, a prolonged residence time of the deposited composition in thelungs may lead to a higher continuous exposure of the active compound atthe site of action. At the same time, it may reduce the requiredfrequency of administration and therefore, enhance patient convenienceand compliance.

In order to achieve a prolonged retention of the active compound ingeneral, various formulation strategies may be pursued, e.g. conversionof the highly water soluble active compound into a less soluble solidform, such as a poorly soluble salt. As a consequence, the compound ispresent in the aerosol in undissolved form, such as in form of a micro-or nanosuspension. Upon deposition of the aerosol droplets, the liquidphase of the composition combines with the physiological fluid, e.g.mucus, and allows the drug to dissolve.

A different formulation strategy is based on the fact that polymericexcipient(s), as described below, may have an effect on the release ofthe active compound from the formulation, and/or on the local residencetime of the composition after deposition onto the target tissue.Therefore, such excipients affect the local bioavailability of theactive compound at the site of action as well.

In one of the preferred embodiments, the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, is formulate with apolymeric excipient to effect slow release and prolonged localretention. Potentially suitable polymers include, in particular,pharmaceutically acceptable water-soluble or water-dispersible polymers,such as methylcellulose, hydroxyethylcellulose, alginate, galactomannan,dextran, agar, guar gum, tragacanth, and mixtures thereof.

If one or several polymeric excipients are present in the liquidcomposition of the invention, care should be taken of the influence onthe dynamic viscosity of such a composition in order to ensure efficientaerosolization. Thus, the dynamic viscosity should not exceed about 1.7mPas. In general, the exact grade of the polymer(s) and the presence ofother excipients should be considered to determine the content of thepolymer(s) in such liquid composition.

It is known that other excipients, namely complexing agents, such ascyclodextrins, di- or multivalent metal salts, such ascalcium-magnesium- or aluminium salts, chelating agents, such asethylenediaminetetraacetic acid including its salts, or amphiphilicagents, such as phospholipids or lecithins, may in a similar mannerprolong the release of the active compound as e.g. polymeric excipients.

The liquid composition of the invention may comprise furtherpharmaceutically acceptable excipients, e.g. osmotic agents, such asinorganic salts; excipients for adjusting and/or buffering the pH, suchas organic or inorganic salts, acids and bases, bulking agents andlyophilisation aids, such as sucrose and lactose, sugar alcohols, likemannitol, sorbitol, and xylitol, stabilizers and antioxidants, such asvitamin E including its derivatives, lycopene including its derivativesand ascorbic acid, ionic and non-ionic surfactants, such asphospholipids and polysorbates, taste-modifying agents, disintegrants,colouring agents, sweeteners, and/or flavours.

In one of the preferred embodiments, one or more osmotic agents, such assodium chloride, are incorporated in the composition to adjust theosmolality to a value in a preferred range as outlined herein above. Ina more preferred embodiment, the osmotic agent is sodium chloride.

In order to provide a well tolerated aerosol, the preparation accordingto the invention should be adjusted to a euhydric pH. The term“euhydric” implies that there may be a difference between pharmaceuticaland physiological requirements so that a compromise has to be foundwhich, for example, ensures that, on one hand, the preparation issufficiently stable during storage, but, on the other hand, is stillwell tolerated. Preferably, the pH value lies in the slightly acidic toneutral region, i.e. between about 4 and about 8. In general, deviationstowards a weakly acidic milieu are tolerated better than an alkalineshift. Particularly preferred is a composition having a pH lying withinthe range from about 4.5 and about 7.5.

For adjusting the pH of the composition of the invention and/orbuffering such composition, physiologically acceptable acids, bases,salts, and combination of these may be used. Suitable excipients forlowering the pH value and/or as acidic components of a buffer system arestrong mineral acids, such as sulphuric acid and hydrochloric acid.Inorganic and organic acids of medium strength, such as phosphoric acid,citric acid, tartaric acid, succinic acid, fumaric acid, methionine,lactic acid, acetic acid, glucuronic acid, as well as acidic salts, suchas hydrogen phosphates with sodium or potassium, may be used as well.Suitable excipients for raising the pH value and/or as basic componentsof a buffer system are mineral bases, such as sodium hydroxide, or otheralkali and alkaline earth hydroxides and oxides, such as magnesiumhydroxide, calcium hydroxide, or basic ammonium salts, such as ammoniumhydroxide, ammonium acetate, or basic amino acids, such as lysine, orcarbonates, such as sodium or magnesium carbonate, sodium hydrogencarbonate, or citrates, such as sodium citrate.

In a preferred embodiment, the composition of the invention comprises atleast one excipient to adjust the pH. In a more preferred embodiment,that excipient is sodium hydroxide.

Mainly for pharmaceutical reasons the chemical stabilisation of thecomposition of the invention by further additives may be indicated. Themost common degradation reactions of a chemically defined activecompound in aqueous preparations comprise, in particular, hydrolysisreactions which may be limited primarily by optimal pH adjustment, aswell as oxidation reactions. As the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)comprises a lysine residue having a primary amino group, the latter, forexample, may be subject to oxidative attack. Therefore, the addition ofan antioxidant, or an antioxidant in combination with a synergist, maybe advisable or necessary.

Antioxidants are natural or synthetic substances which are capable ofpreventing or inhibiting the oxidation of the active compound.Antioxidants are primarily ajuvants which are oxidizable and/or act asreducing agents, such as tocopherol acetate, lycopene, reducedglutathione, catalase, peroxide dismutase. Further suitable antioxidantsare, for example, ascorbic acid, sodium ascorbate and other salts andderivatives of ascorbic acid, e.g. ascorbyl palmitate, fumaric acid andits salts, malic acid and its salts.

Synergistic substances are those which do not directly act as reactantsin oxidation processes, but which counteract such processes by indirectmechanisms, for example, by complexation of metal ions which are knownto act catalytically in oxidation processes. Ethylenediaminetetraaceticacid (EDTA) and salts and derivatives thereof, citric acid and saltsthereof, malic acid and salts thereof, are such synergistic substanceswhich may act as chelating agents.

In one of the embodiments, the composition of the invention comprises atleast one antioxidant. In a further embodiment, the compositioncomprises both an antioxidant and a chelating agent.

As mentioned above, the composition of the invention may comprise anexcipient affecting the taste. A bad taste is extremely unpleasant andirritating, especially in inhalation administration, and can result innon-compliance, and thus, therapy failure. The bad taste is perceived bythe patient through that part of the aerosol which precipitates in theoral and pharyngeal region during inhalation. Even if the particle sizeof the aerosol can be optimized in such a manner that only a smallfraction of the preparation precipitates in the above mentioned regions(said fraction being lost for therapy, unless the oral, pharyngeal ornasal mucosa is the target tissue) it is presently hardly possible toreduce said fraction to such an extent that the bad taste of an activecompound is no longer perceived. Therefore, the improvement of the tasteof a composition or the masking of the taste of an active compound maybe crucial.

In order to improve the taste of the composition, one or morepotentially useful excipients from the group of sugars, sugar alcohols,salts, flavours, complexing agents, polymers, sweeteners, such as sodiumsaccharin, aspartame, surfactants may be incorporated.

In a preferred embodiment, the composition of the invention comprises atleast one taste-modifying excipient. In a more preferred embodiment,said taste-modifying excipient is sodium saccharin.

In another embodiment, the composition comprises a further activecompound, the combination of which with the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) havinga combined or, ideally, a synergistic therapeutic effect.

In case a liquid formulation for aerosolization may not have asufficiently long shelf life to serve as a suitable formulation formarket it may be beneficial to provide a solid composition instead. Suchsolid composition generally has the potential for a longer shelf lifecompared to a liquid composition.

The solid composition of the invention comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and at least oneexcipient. In general, the same excipients as already described abovemay be selected. Depending on the manufacturing process/method of thesolid composition one or more additional excipients may be useful. Ifthe solid composition is, for example, prepared by freeze drying(lyophilization) being one of the preferred methods of preparing such asolid composition, it may be advantageous to incorporate at least onebulking agent and/or lyophilization aid, e.g. a sugar, such as sucrose,fructose, glucose, trehalose, or a sugar alcohol, such as mannitol,sorbitol, xylitol, isomalt.

The solid composition is further characterized in that it is dissolvableor dispersible in an aqueous liquid solvent. As defined herein in thedescription and the claims, the term “dissolvable” means that the solidcomposition and the aqueous liquid solvent can be combined to form asolution or colloidal solution, whereas the term “dispersible” should beinterpreted to include the formation of liquid dispersions, inparticular, emulsions and microsuspensions. The term “aqueous” meansthat the major liquid constituent of the solvent is water. Solvents andco-solvents other than water should be avoided. In another embodiment,the aqueous liquid solvent comprises at least about 80 wt.-% of water.In yet another embodiment, at least about 90 wt.-% of the liquidconstituents of the solvent is water. If the incorporation of anon-aqueous solvent, such as ethanol, glycerol, propylene glycol orpolyethylene glycol, cannot be avoided, the excipient should be selectedcarefully and in consideration of its physiological acceptability andthe therapeutic use of the composition. According to a preferredembodiment, the composition is substantially free of non-aqueoussolvents. If the incorporation of a non-aqueous solvent, such asethanol, glycerol, propylene glycol or polyethylene glycol, cannot beavoided, the same precautions have to be considered as described above.According to another preferred embodiment, the aqueous solvent todissolve or disperse the solid composition is substantially free ofnon-aqueous solvents.

The amount of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in the solid compositionhas to correspond to a concentration within a range from about 4 mg/mLto about 100 mg/mL following dissolution or dispersion in an aqueousliquid solvent. More preferably, the amount of the above activecompound, or any pharmaceutically acceptable salt thereof, has tocorrespond to a concentration within the range from about 17 mg/mL toabout 95 mg/mL, or about 35 mg/mL to about 95 mg/mL, respectively, or,even more preferred, about 70 mg/mL to about 95 mg/mL, followingdissolution or dispersion in an aqueous liquid solvent.

In a further embodiment, the counter ion of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) in thesolid composition is acetate.

The solid composition of the invention for reconstitution may be part ofa pharmaceutical kit. Such kit preferably comprises the solidcomposition in sterile form. As used herein in the description and theclaims the terms “sterile” or “sterility” are defined according to theusual pharmaceutical meaning and thus to be understood as the absence ofgerms which are capable of reproduction. Sterility is determined withsuitable tests which are defined in the relevant pharmacopoeias.According to current scientific standards, a sterility assurance level(SAL) of 10⁻⁶, i.e. assurance of less than one chance in one millionthat viable micro-organisms are present in a sterilized article), isregarded as acceptable for sterile preparations. In practice,contamination rates may be higher, and contamination rates foraseptically manufactured preparations might amount to about 10⁻³. Forpractical reasons, it remains difficult to demand sterility in anabsolute sense. Therefore, the sterility of the composition of theinvention should be understood herein in the description and the claimsin such manner that said composition meets the requirements of therelevant pharmacopeia with respect to sterility.

The solid composition of the invention may be prepared by providing aliquid composition being similar to the liquid composition ready foraerosolization and subsequently drying it, e.g. by lyophilisation.Similar means that the liquid composition from which the solidcomposition is prepared by drying may not comprise all solid ingredientsof the ready-to-go liquid composition, for example, in the case that theliquid carrier for reconstitution is designed to comprise one or more ofthe excipients. It is even not necessary, that the concentrations of theingredients of these two liquid compositions are identical. The solidcomposition of the invention may even be prepared, for example, byproviding the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and, optionally, at leastone excipient, in powder form and subsequently mixing such powder with apowder comprising the remaining excipients to finally form a powdermixture.

In another aspect, the invention provides a pharmaceutical kit for thepreparation and delivery of a pharmaceutical aerosol for pulmonaryadministration comprising a dispersed liquid phase and a continuous gasphase, wherein the dispersed liquid phase comprises aqueous dropletscomprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, has a mass median diameterfrom about 1.5 μm to about 5 μm, and has a droplet size distributionhaving a geometrical standard deviation from about 1.2 to about 1.7. Thekit is further characterized by comprising an aerosol nebulizer and ancomposition comprising a concentration within a range from about 4 mg/mLto about 100 mg/mL of the active compound, or any pharmaceuticallyacceptable salt thereof; or comprising a nebulizer and a solidpharmaceutical composition for preparing the liquid composition, whereinthe composition comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and wherein the solidcomposition is dissolvable or dispersible in an aqueous liquid solvent,and wherein the liquid composition comprises a concentration within arange from about 4 mg/mL to about 100 mg/mL of the active compound, orany pharmaceutically acceptable salt thereof.

Nebulizers are devices capable of aerosolizing liquids. Preferably, thenebulizer of the kit of the invention is selected from jet nebulizers,ultrasonic nebulizers, piezoelectronic nebulizers, jet collisionnebulizers, electrohydrodynamic nebulizers, capillary force nebulizers,perforated membrane nebulizers and perforated vibrating membranenebulizers (M. Knoch, M. Keller, Expert Opin. Drug Deliv., 2005, 2,377). Particularly preferred are piezoelectric, electrohydrodynamicand/or perforated membrane-type nebulizers, e.g. nebulizers from thedrug delivery platforms Mystic™ (Battelle Pharma [Battelle MemorialInstitute], United States), eFlow™ (Pari GmbH, Germany), Aeroneb™,Aeroneb Pro™, Aero Dose™ (Aerogen Inc, United States). These types ofnebulizers are particularly useful if the aerosol is to be delivered tothe bronchi and/or lungs.

Preferably, the nebulizer should be selected or adapted to be capable ofaerosolizing the liquid composition at a rate of at least about 0.1mL/min. More preferably, the nebulizer is capable of an (total) outputrate (the rate at which the aerosol is emitted from the aerosolgenerator) of at least about 0.150 mL/min or at least about 150 mg/minfor those liquid compositions the densities of which are—for practicalpurposes—close to 1 g/mL, i.e. within the range from about 0.95 g/mL toabout 1.05 g/mL. In further embodiments, the output rate of thenebulizer is within the range from about 200 mg/min to about 700 mg/min,or from about 250 mg/min to about 650 mg/min, respectively.

The nebulizer should also preferably be selected or adapted to becapable of aerosolizing and emitting the liquid composition at a meandelivery rate of at least about 0.8 mg of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, per minute. The (mean)delivery rate of a drug or active compound is a parameter to determinethe amount of drug or active compound a patient might be expected toreceive during a treatment period. In further embodiments, the nebulizeris selected or adapted to enable a mean delivery rate of the activecompoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, at the range from about 3mg per minute to about 25 mg per minute or at the range from about 5 mgper minute to about 18 mg per minute, respectively.

According to a further preference, the nebulizer should be selected oradapted to be capable of aerosolizing and emitting at least of about 70wt.-% of the loaded dose of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, whereas said fraction ofthe loaded dose is comprised of droplets having a mass median diameterof not more than about 5 μm. A fraction of a dispersed phase having adroplet size of not more than about 5 μm is often referred to as therespirable fraction, as droplets of said size—in contrast to largerdroplets—have a high chance of being deposited in the lungs, instead ofthe trachea and the pharynx. More preferably, at least of about 80 wt.-%of the dose filled into the nebulizer is aerosolized to droplets of asize of not more than about 5 μm and emitted from the device. Such adevice may be best selected by using an, optionally customized,electronic nebulizer based on the vibrating perforated membrane design,such as a nebulizer from the eFlow™ drug delivery platform (Pari GmbH,Germany). According to even more preferred embodiments, a least of about85 wt.-% and about 90 wt.-%, respectively, of the loaded dose isaerosolized to droplets of a size of not more than about 5 μm andemitted.

In another aspect the invention provides a method of preparing anddelivering an aerosol for pulmonary administration, said methodcomprising the steps of providing a liquid pharmaceutical compositioncomprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in a concentration withina range from about 4 mg/mL to about 100 mg/mL, or providing a solidpharmaceutical composition for preparing the liquid composition, whereinthe composition comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, and wherein the solidcomposition is dissolvable or dispersible in an aqueous liquid solvent,and wherein the liquid composition comprises a concentration within arange from about 4 mg/mL to about 100 mg/mL of the active compound, orany pharmaceutically acceptable salt thereof, and providing a nebulizercapable of aerosolizing said liquid pharmaceutical composition at a meandelivery rate of at least about 0.8 mg of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, per minute, the nebulizerbeing further adapted to emit an aerosol comprising a dispersed liquidphase having a mass median diameter from about 1.5 μm to about 5 μm, andhaving a droplet size distribution having a geometrical standarddeviation from about 1.2 to about 1.7, and operating the nebulizer toaerosolize the liquid pharmaceutical composition.

The composition of the invention, whether liquid, initially solid orfinally aerosolized, or the pharmaceutical kit comprising thecomposition, can be used for the prevention, management or treatment ofdiseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity, e.g. pulmonary diseases, such asalpha-1 antitrypsin deficiency (AATD), cystic fibrosis (CF), non-cysticfibrosis bronchiactasis (NCFB), or chronic obstructive pulmonary disease(COPD), or infections of the lungs causing diseases or conditions of thelungs, being mediated by human neutrophil elastase activity.

As used herein, the term “prevention”/“preventing” e.g. preventivetreatments comprise prophylactic treatments. In preventive applications,the pharmaceutical composition or the pharmaceutical aerosol of theinvention is administered to a subject suspected of having, or at riskfor developing diseases or conditions of the lungs being mediated by orresulting from human neutrophil elastase activity.

As used herein, the term “management” means increasing the time toappearance of a symptom of diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity or amark associated with diseases or conditions of the lungs being mediatedby or resulting from human neutrophil elastase activity or slowing theincrease in severity of a symptom of diseases or conditions of the lungsbeing mediated by or resulting from human neutrophil elastase activity.Further, “management” as used herein includes reversing or inhibition ofdisease progression. “Inhibition” of disease progression or diseasecomplication in a subject means preventing or reducing the diseaseprogression and/or disease complication in the subject.

The terms “treatment”/“treating” as used herein includes: (1) delayingthe appearance of clinical symptoms of the state, disease or conditiondeveloping in an animal, particularly a mammal and especially a human,that may be afflicted with or predisposed to the state, disease orcondition but does not yet experience or display clinical or subclinicalsymptoms of the state, disease or condition; (2) inhibiting the state orcondition (e.g. arresting, reducing or delaying the development of thedisease, or a relapse thereof in case of maintenance treatment, of atleast one clinical or subclinical symptom thereof; and/or (3) relievingthe condition (i.e. causing regression of the state, disease orcondition or at least one of its clinical or subclinical symptoms). Thebenefit to a patient to be treated is either statistically significantor at least perceptible to the patient or to the physician. However, itwill be appreciated that when a medicament is administered to a patientto treat a disease, the outcome may not always be effective treatment.

In therapeutic applications, the pharmaceutical composition is usuallyadministered to a subject such as a patient already suffering fromdiseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity, in an amount sufficient to cure orat least partially arrest the symptoms of the disease or condition.Amounts effective for this use will depend on the severity and course ofthe disease, previous therapy, the subject's health status and responseto the drugs, and the judgment of the treating physician.

In the case wherein the subject's condition does not improve, thepharmaceutical composition or the pharmaceutical aerosol of theinvention may be administered chronically, which is, for an extendedperiod of time, including throughout the duration of the subject's lifein order to ameliorate or otherwise control or limit the symptoms of thesubject's disease or condition.

In the case wherein the subject's status does improve, thepharmaceutical composition or the pharmaceutical aerosol may beadministered continuously; alternatively, the dose of drugs beingadministered may be temporarily reduced or temporarily suspended for acertain length of time (i.e., a “drug holiday”).

Once improvement of the patient's condition has occurred, a maintenancedose of the pharmaceutical composition or the pharmaceutical aerosol ofthe invention is administered if necessary. Subsequently, the dosage orthe frequency of administration, or both, is optionally reduced, as afunction of the symptoms, to a level at which the improved disease isretained.

Thus in another aspect the invention provides a pharmaceuticalcomposition comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof; wherein OctG is(S)-2-aminodecanoic acid; ^(D)Pro is D-proline; and optionally one ormore pharmaceutically acceptable diluents, excipients or carriers, foruse in a method for the prevention, management or treatment of diseasesor conditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject, preferably for use in amethod for treatment of diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity in asubject.

Also provided is the use of the pharmaceutical composition as describedherein for the manufacture of a medicament for the prevention,management or treatment of diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity in asubject, preferably for the manufacture of a medicament for thetreatment of diseases or conditions of the lungs being mediated by orresulting from human neutrophil elastase activity in a subject.

Also provided is the use of a pharmaceutical composition as describedherein for the prevention, management or treatment of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject, preferably for the treatmentof diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity in a subject.

Also provided is a method for the prevention, management or treatment ofdiseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity in a subject, preferably a method forthe treatment of diseases or conditions of the lungs being mediated byor resulting from human neutrophil elastase activity in a subject,comprising administering to said subject a pharmaceutical composition asdescribed herein e.g. administering to said subject a therapeuticallyeffective amount of a pharmaceutical composition as described herein.

The term “pharmaceutically acceptable diluent, excipient or carrier” asused herein refers to a carrier or excipient or diluent that is suitablefor use with humans and/or animals without undue adverse side effects(such as toxicity, irritation, and allergic response) commensurate witha reasonable benefit/risk ratio. It can be a pharmaceutically acceptablesolvent, suspending agent or vehicle, for delivering the instantcompounds to the subject.

Determination of a therapeutically effective amount is well within thecapability of those skilled in the art, especially in light of thedetailed disclosure provided herein. In some embodiments, atherapeutically effective amount of the active compound or apharmaceutically acceptable salt thereof, may (i) reduce theconcentration of active elastase in sputum of a subject, ii) may inhibitthe activity of human neutrophil elastase activity in sputum of asubject in various embodiments, the amount is sufficient to ameliorate,palliate, lessen, and/or delay one or more of symptoms of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject.

The therapeutically effective amount may vary depending on the subject,and disease or condition being treated, the weight and age of thesubject, the severity of the disease or condition, and the manner ofadministering, which can readily be determined by one ordinary skilledin the art.

In one embodiment the diseases or conditions of the lungs being mediatedby or resulting from human neutrophil elastase activity are pulmonarydiseases such as alpha-1 antitrypsin deficiency (AATD), cystic fibrosis(CF), non-cystic fibrosis bronchiactasis (NCFB), or chronic obstructivepulmonary disease (COPD), or infections of the lungs causing diseases orconditions of the lungs, being mediated by human neutrophil elastaseactivity.

In a preferred embodiment the diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity arepulmonary diseases, wherein the pulmonary disease is non-cystic fibrosisbronchiactasis (NCFB) or cystic fibrosis (CF).

In a more preferred embodiment the diseases or conditions of the lungsbeing mediated by or resulting from human neutrophil elastase activityare pulmonary diseases, wherein the pulmonary disease is cystic fibrosis(CF).

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, is administered to thesubject as pharmaceutical aerosol for pulmonary administrationcomprising a dispersed liquid phase and a continuous gas phase, whereinthe dispersed liquid phase

-   -   (a) comprises aqueous droplets comprising the active compound        cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-);    -   or any pharmaceutically acceptable salt thereof; wherein    -   OctG is (S)-2-aminodecanoic acid;    -   ^(D)Pro is D-proline;    -   (b) has a mass median diameter from about 1.5 μm to about 5 μm;        and    -   (c) has a droplet size distribution having a geometrical        standard deviation from about 1.2 to about 1.7.

Preferably the aerosol being emitted from an aerosol generator at a rateof at least about 0.1 mL dispersed liquid phase per minute.

Equally preferably the aerosol being emitted from an aerosol generatorat a mean delivery rate of at least about 0.8 mg of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; per minute.

The aerosol is preferably emitted from an aerosol generator at a rateand at a mean delivery rate as described in the preferred embodimentsabove.

In one embodiment the pharmaceutical composition for use in a method forthe prevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject, is a liquid pharmaceutical composition forpreparing an aerosol as described herein, wherein the liquidpharmaceutical composition comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, in a concentration withina range from about 4 mg/mL to about 100 mg/mL, preferably, within arange from about 17 mg/mL to about 95 mg/mL, or about 35 mg/mL to about95 mg/mL, respectively, and more preferably, within a range from about70 mg/mL to about 95 mg/mL.

In another aspect the invention provides a pharmaceutical aerosol forpulmonary administration comprising a dispersed liquid phase and acontinuous gas phase, wherein the dispersed liquid phase

-   -   (a) comprises aqueous droplets comprising the active compound        cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-);        or any pharmaceutically acceptable salt thereof; wherein    -   OctG is (S)-2-aminodecanoic acid;    -   ^(D)Pro is D-proline;    -   (b) has a mass median diameter from about 1.5 μm to about 5 μm;        and    -   (c) has a droplet size distribution having a geometrical        standard deviation from about 1.2 to about 1.7, for use in a        method for the prevention, management or treatment of diseases        or conditions of the lungs being mediated by or resulting from        human neutrophil elastase activity in a subject.

The aerosol is preferably emitted from an aerosol generator at a rateand at a mean delivery rate as described in the preferred embodimentsabove.

Also provided is the use of the pharmaceutical aerosol as describedherein for the manufacture of a medicament for the prevention,management or treatment of diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity in asubject, preferably for the manufacture of a medicament for thetreatment of diseases or conditions of the lungs being mediated by orresulting from human neutrophil elastase activity in a subject.

Also provided is the use of the pharmaceutical aerosol as describedherein for the prevention, management or treatment of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject, preferably for the treatmentof diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity in a subject.

Also provided is a method for the prevention, management or treatment ofdiseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity in a subject, preferably a method forthe treatment of diseases or conditions of the lungs being mediated byor resulting from human neutrophil elastase activity in a subject,comprising administering to said subject the pharmaceutical aerosol asdescribed herein e.g. administering to said subject a therapeuticallyeffective amount of a pharmaceutical aerosol as described herein.

In one embodiment the diseases or conditions of the lungs being mediatedby or resulting from human neutrophil elastase activity are pulmonarydiseases such as alpha-1 antitrypsin deficiency (AATD), cystic fibrosis(CF), non-cystic fibrosis bronchiactasis (NCFB), or chronic obstructivepulmonary disease (COPD), or infections of the lungs causing diseases orconditions of the lungs, being mediated by human neutrophil elastaseactivity.

In a preferred embodiment the diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity arepulmonary diseases, wherein the pulmonary disease is non-cystic fibrosisbronchiactasis (NCFB) or cystic fibrosis (CF).

In a more preferred embodiment the diseases or conditions of the lungsbeing mediated by or resulting from human neutrophil elastase activityare pulmonary diseases, wherein the pulmonary disease is cystic fibrosis(CF).

The counter ion of the active compound of the pharmaceutical compositionor the pharmaceutical aerosol for use in a method for the prevention,management or treatment of diseases or conditions of the lungs beingmediated by or resulting from human neutrophil elastase activity in asubject is as described for the active compound above and is preferablyacetate.

The pharmaceutical composition or the pharmaceutical aerosol for use ina method for the prevention, management or treatment of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject is usually administered to thesubject by oral inhalation or intratracheal, preferably by oralinhalation.

The dosing regimen of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, comprised by thepharmaceutical composition or the pharmaceutical aerosol, in the methodsprovided herein may vary depending upon the indication, route ofadministration, and severity of the condition, for example. Depending onthe route of administration, a suitable dose can be calculated accordingto body weight, body surface area, or organ size. Additional factorsthat can be taken into account include time and frequency ofadministration, drug combinations, reaction sensitivities, andtolerance/response to therapy. The amount, e.g. the therapeuticallyeffective amount of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), or apharmaceutically acceptable salt thereof, may be provided in a singledose or multiple doses to achieve the desired treatment endpoint.

The frequency of dosing will depend on the pharmacokinetic parameters ofthe active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, administered, the route ofadministration, and the particular disease treated. The dose andfrequency of dosing may also depend on pharmacokinetic andpharmacodynamic, as well as toxicity and therapeutic efficiency data.For example, pharmacokinetic and pharmacodynamic information about theactive compound or a pharmaceutically acceptable salt thereof, can becollected through preclinical in vitro and in vivo studies, laterconfirmed in humans during the course of clinical trials. Thus, for theactive compound or a pharmaceutically acceptable salt thereof, used inthe methods provided herein, a therapeutically effective dose can beestimated initially from biochemical and/or cell-based assays. Then,dosage can be formulated in animal models to achieve a desirablecirculating concentration range. As human studies are conducted furtherinformation will emerge regarding the appropriate dosage levels andduration of treatment for various diseases and conditions.

Toxicity and therapeutic efficacy of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, can be determined bystandard pharmaceutical procedures in cell cultures or experimentalanimals, e.g., for determining the LD₅₀ (the dose lethal to 50% of thepopulation) and the ED₅₀ (the dose therapeutically effective in 50% ofthe population). The dose ratio between toxic and therapeutic effects isthe “therapeutic index”, which typically is expressed as the ratioLD₅₀/ED₅₀. Compounds that exhibit large therapeutic indices, i.e., thetoxic dose is substantially higher than the effective dose, arepreferred. The data obtained from such cell culture assays andadditional animal studies can be used in formulating a range of dosagefor human use. The doses of such compounds lies preferably within arange of circulating concentrations that include the ED₅₀ with little orno toxicity.

An exemplary treatment regime entails administration once daily, twicedaily, three times daily, every day, every second day, every third day,every fourth day, every fifth day, every sixth day, twice per week, onceper week. The active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, is usually administered onmultiple occasions. Intervals between single dosages can be, forexample, less than a day, a day, two days, three days, four days, fivedays, six days or a week. The combination of the invention may be givenas a continuous uninterrupted treatment. The combination of theinvention may also be given in a regime in which the subject receivescycles of treatment (administration cycles) interrupted by a drugholiday or period of non-treatment.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose betweenabout 0.1 and about 10000 mg/day.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is is administered to the subject at a dosebetween about 0.001 and about 100 mg/kg.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose betweenabout 5 and about 1000 mg/day.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose betweenabout 20 and about 960 mg/day.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose betweenabout 80 and about 320 mg/day.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose of about20, about 60, about 120, about 240, about 480 or about 960 mg/day.

In one embodiment the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, of the pharmaceuticalcomposition or the pharmaceutical aerosol for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is administered to the subject at a dose of about80, about 160, or about 320 mg/day.

In another aspect the invention provides a kit for the preparation anddelivery of a pharmaceutical aerosol for pulmonary administrationcomprising a dispersed liquid phase and a continuous gas phase, whereinthe dispersed liquid phase

-   -   (a) comprises aqueous droplets comprising the active compound        cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-);    -   or any pharmaceutically acceptable salt thereof; wherein    -   OctG is (S)-2-aminodecanoic acid;    -   ^(D)Pro is D-proline;    -   (b) has a mass median diameter from about 1.5 μm to about 5 μm;        and    -   (c) has a droplet size distribution having a geometrical        standard deviation from about 1.2 to about 1.7;    -   and wherein the kit comprises a nebulizer and a liquid        composition comprising a concentration within a range from about        4 mg/mL to about 100 mg/mL of the active compound; or any        pharmaceutically acceptable salt thereof;    -   or comprises a nebulizer and a solid pharmaceutical composition        for preparing the liquid composition, wherein the composition        comprises the active compound        cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-);        or any pharmaceutically acceptable salt thereof; and wherein the        solid composition is dissolvable or dispersible in an aqueous        liquid solvent, and wherein the liquid composition comprises a        concentration within a range from about 4 mg/mL to about 100        mg/mL of the active compound; or any pharmaceutically acceptable        salt thereof, for use in a method for the prevention, management        or treatment of diseases or conditions of the lungs being        mediated by or resulting from human neutrophil elastase activity        in a subject.

Also provided is the use of the kit as described herein for themanufacture of a medicament for the prevention, management or treatmentof diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity in a subject, preferably for themanufacture of a medicament for the treatment of diseases or conditionsof the lungs being mediated by or resulting from human neutrophilelastase activity in a subject.

Also provided is the use of the kit as described herein for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject, preferably for the treatment of diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity in a subject.

Also provided is a method for the prevention, management or treatment ofdiseases or conditions of the lungs being mediated by or resulting fromhuman neutrophil elastase activity in a subject, preferably a method forthe treatment of diseases or conditions of the lungs being mediated byor resulting from human neutrophil elastase activity in a subject,comprising administering to said subject the pharmaceutical aerosol ofthe kit as described herein e.g. administering to said subject atherapeutically effective amount of a pharmaceutical aerosol of the kitas described herein.

In one embodiment the nebulizer of the kit for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is selected from the group consisting of jetnebulizers, ultrasonic nebulizers, piezoelectronic nebulizers, jetcollision nebulizers, electrohydrodynamic nebulizers, capillary forcenebulizers, perforated membrane nebulizers and perforated vibratingmembrane nebulizers.

In one embodiment the nebulizer of the kit for use in a method for theprevention, management or treatment of diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity in a subject is adapted to be capable of aerosolizing theliquid composition at a rate of at least about 0.8 mg of the activecompoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; per minute.

In one embodiment at least about 70 wt.-% of the loaded dose of theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, comprised by the kit iscomprised of droplets having a mass median diameter of not more thanabout 5 μm.

In one embodiment the counter ion of the active compound comprised bythe kit is as described for the active compound above and is preferablyacetate.

In another aspect the invention provides a kit comprising the activecompoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany

-   -   pharmaceutically acceptable salt thereof; wherein    -   OctG is (S)-2-aminodecanoic acid;    -   ^(D)Pro is D-proline;    -   and a package insert wherein the package insert comprises        instructions for treating a subject for diseases or conditions        of the lungs being mediated by or resulting from human        neutrophil elastase activity using the active compound.

In one embodiment the kit comprises the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, as aerosol for pulmonaryadministration comprising a dispersed liquid phase and a continuous gasphase, wherein the dispersed liquid phase

-   -   (a) comprises aqueous droplets comprising the active compound        cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-);        or any pharmaceutically acceptable salt thereof;    -   (b) has a mass median diameter from about 1.5 μm to about 5 μm;        and    -   (c) has a droplet size distribution having a geometrical        standard deviation from about 1.2 to about 1.7.

In one embodiment at least about 70 wt.-% of the loaded dose of theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-), orany pharmaceutically acceptable salt thereof, comprised by the kit iscomprised of droplets having a mass median diameter of not more thanabout 5 μm.

In one embodiment the counter ion of the active compound comprised bythe kit is as described for the active compound above and is preferablyacetate.

The following Examples illustrate the present invention but are not tobe construed as limiting its scope in any way.

EXAMPLES Example 1

248.18 g of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) basedon a net peptide content of 93.03% (see calculation below) andcorresponding to 230.88 g of net peptide, were dissolved in 0.5% (w/w)aqueous sodium chloride solution. The solution was adjusted to pH 5.5with 154 g 1 M sodium hydroxide and finally 0.5% (w/w) aqueous sodiumchloride solution was added to a total weight of 2957 g. After sterilefiltration through 2×0.22 μm pore size filters the product was packed inPh.Eur. Type 1 glass vials with fluoropolymer coated bromobutyl rubberstoppers and tear-off plain aluminium overseals. The strength of thesolution was 80 mg/mL.

Calculation of the net peptide content of the drug substance (activecompound):Net peptide content[%]=[(100−impurity[%]/100)×(100−watercontent[%]/100)×(100−residual solvent[%]/100)×(100−residualTFA/100)×freesalt[%]/100]×100=[(100−0.7/100)×(100−2.5/100)×(100−0.013/100)×96.1/100]×100=93.03%

Example 2

The formulation of Example 2 was prepared as described in Example 1except 0.6% (w/w) aqueous sodium chloride solution was used.

Example 3

4.2 g of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)corrected for 95% purity were dissolved in 40 mL 0.6% (w/w) aqueoussodium chloride solution. The solution was adjusted to pH 5.5 with 1 Msodium hydroxide and finally 0.6% (w/w) sodium chloride solution wasadded to a total volume of 50 mL. After sterile filtration under asepticconditions (laminar air flow) using an PES (Polyethersulfone) 0.2 μmsyringe filter the formulation was aliquoted into sterile 5 mL vialswith sterile teflon coated rubber stoppers and stored refrigerated at5±3° C. and room temperature (25±2° C.), respectively. The strength ofthe solution was 80 mg/mL. During 12 weeks the solution was observed forstability. Physicochemical parameters (Table 1) and aerosolcharacteristics (Table 2) of an aerosol prepared and delivered by aneFlow® 30 XL electronic nebulizer (Pari Pharma GmbH, Starnberg, Germany)were determined at the beginning and at the end of the observationperiod. Physicochemical characterization and determination of aerosolcharacteristics (laser diffraction with Malvern Mastersizer X, V2.15,[Malvern Instruments GmbH, Herrenberg, Germany]), were performed by PariPharma GmbH, BU Pharma, Gräfeling, Germany, according topharmacopoeia-compliant methods.

TABLE 1 Physiochemical properties of formulation of Example 3 AfterAfter 12 weeks 12 weeks Example 3 Initial at 5° C. at 25° C. pH 5.5 5.45.4 Osmolality [mOsmol/kg] 353 355 356 Viscosity [mPa * s] 1.44 1.411.41 Surface tension [mN/m] 50.0 50.3 50.2

The physicochemical properties of the above formulation remain unchangedduring 12 weeks at 5° C. and 25° C., respectively.

TABLE 2 Aerosol characteristics of formulation of Example 3, determinedin triplicate After After 12 weeks 12 weeks Example 3 Initial at 5° C.at 25° C. Mass median diameter [μm] 3.00 ± 0.12 2.89 ± 0.05 2.92 ± 0.04Geometric standard deviation 1.55 ± 0.03 1.53 ± 0.01 1.54 ± 0.01Respirable fraction <5 μm [%] 86.18 ± 2.52  88.60 ± 1.14  78.91 ± 0.61 Total output rate [mg/min] 338 ± 25  304 ± 9  320 ± 10 

During storage of 12 weeks at 5° C. and 25° C., respectively, nosignificant (P=95%, n=3) changes in mass median diameter, geometricstandard deviation and respirable fraction (<5 μm) were observed. Onlythe total output rate of the 5° C. sample was slightly decreased.

Example 4

1.05 g of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)corrected for 95% purity were dissolved in 0.9% (w/w) aqueous sodiumchloride solution to obtain a total volume of 10 mL. The strength of thesolution was 100 mg/mL. Physicochemical parameters (Table 3) and aerosolcharacteristics (Table 4) of an aerosol prepared and delivered by aneFlow® 30 XL electronic nebulizer were determined by Pari Pharma GmbH,BU Pharma, Gräfeling, Germany, as explained above.

TABLE 3 Physiochemical properties of formulation of Example 4 Example 4pH 4.32 Osmolality [mOsmol/kg] 433 Viscosity [mPa * s] 1.62 Surfacetension [mN/m] 49.8

TABLE 4 Aerosol characteristics of formulation of Example 4, determinedin triplicate Example 4 Mass median diameter [μm]  3.31 ± 0.10 Geometricstandard deviation  1.55 ± 0.02 Respirable fraction <5 μm [%] 81.75 ±2.33 Total output rate [mg/min] 549.7 ± 21.3

Example 5

1.05 g of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)corrected for 95% purity were dissolved in 0.9% (w/w) aqueous sodiumchloride solution to obtain a total volume of 10 mL. 0.02% (w/w)Polysorbate 80 were added subsequently. The strength of the solution was100 mg/mL. Physicochemical parameters (Table 5) and aerosolcharacteristics (Table 6) of an aerosol prepared and delivered by aneFlow® 30 XL electronic nebulizer were determined by Pari Pharma GmbH,BU Pharma, Gräfeling, Germany, as explained above.

TABLE 5 Physiochemical properties of formulation of Example 5 Example 5pH 4.44 Osmolality [mOsmol/kg] 435 Viscosity [mPa * s] 1.55 Surfacetension [mN/m] 48.6

TABLE 6 Aerosol characteristics of formulation of Example 5, determinedin triplicate Example 5 Mass median diameter [μm] 2.77 ± 0.02 Geometricstandard deviation 1.51 ± 0.01 Respirable fraction <5 μm [%] 91.19 ±0.56  Total output rate [mg/min] 273.7 ± 3.5 

Example 6a-e

5.4 g of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) basedon a net peptide content of 93.03% (see calculation above) andcorresponding to 5.023 g of net peptide, were dissolved in 0.5% (w/w)aqueous sodium chloride solution. The solution was adjusted to pH 5.5with 1 M sodium hydroxide and finally 0.5% (w/w) sodium chloridesolution was added to a total volume of 71.8 mL. The strength of thesolution was 70 mg/mL (Example 6a).

Diluted solutions with strengths of 35 mg/mL (Example 6b), 17.4 mg/mL(Example 6c), 8.8 mg/mL (Example 6d) and 4.3 mg/mL (Example 6e) wereprepared with placebo (0.5% [w/w] aqueous sodium chloride solution)according to Table 7.

TABLE 7 Preparation scheme for diluted solutions of Examples 6b, 6c, 6dand 6e Aliqot of 70 mg/mL Final strength pH formulation Amount ofplacebo of diluted solution of diluted [mL] [mL] [mg/mL] solutions^(a))10 10 35 5.46 10 30 17.4 5.43 2.5 17.5 8.8 5.42 2.5 37.5 4.3 5.40^(a))The pH of the placebo used for the dilution step was 4.76 and thepH of the corresponding diluted solution was not re-adjusted to pH 5.5.

Example 6f

The formulation of Example 6f was prepared according to the proceduredescribed in Example 1. The strength of the solution was 80 mg/mL.

Determination of Delivery Rates (Mean) and Total Delivered Doses ofExamples 6a, 6c, 6e and 6f

The determination of delivery rates and total delivered doses wasperformed by Intertek Melbourn Scientific (Melbourn, UK) in triplicatefor 70 mg/mL (Example 6a), 17.4 mg/mL (Example 6c) and 4.3 mg/mL(Example 6e) formulations, and in quintuplicate for the 80 mg/mL(Example 6f) formulation using Pari eFlow® XL 30 devices (Pari Pharma,Starnberg, Germany) and a suitable, pharmacopoeia-compliant method forsaid determination of formulations comprising acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-).

The results (mean values) are summarized in Table 8.

TABLE 8 Delivery rates and total delivered doses of Examples 6a, 6c, 6eand 6f Example 6f 6a 6c 6e Strength of 80 70 17.4 4.3 solution [mg/mL]Delivered mass 0.7566 3.7202 3.7402 3.7928 (mean) [g] Mean delivery 8.08.1 3.2 0.8 rate [mg/min] Mean of total 32.1 131.7 25.9 8.2 deliveredactive compound [mg] Mean efficiency^(a)) 53.0 50.6 39.8 50.3 [%]^(a))Mean efficiency [%] is mean of total delivered active compound(dose, actual)/mean of theoretical dose delivered (calculated by usingdelivered mass (mean) and strength of solution, density assumed to be 1g/mL)

Aerodynamic Particle Size Distribution (APSD) Determination of Examples6a, 6c, 6e and 6f

The APSD determination was performed by Intertek Melbourn Scientific(Melbourn, UK) in triplicate for 70 mg/mL (Example 6a), 17.4 mg/mL(Example 6c) and 4.3 mg/mL (Example 6e) formulations, and inquintuplicate for the 80 mg/mL (Example 6f) formulation using the NextGeneration Impactor (NGI), Pari eFlow® XL 30 devices (Pari Pharma,Starnberg, Germany) and a suitable, pharmacopoeia-compliant method forthe APSD of formulations comprising acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-).

The results (mean values) are summarized in Table 9.

TABLE 9 Aerodynamic particle size distribution determination (APSD) ofExamples 6a, 6c, 6e and 6f Example 6f 6a 6c 6e Strength of 80 70 17.44.3 solution [mg/mL] NGI Stages Throat [mg] 0.62 1.39 0.50 0.12 Stage 1[mg] 0.90 3.20 1.01 0.24 Stage 2 [mg] 0.80 2.98 1.02 0.24 Stage 3 [mg]4.80 21.44 7.14 1.82 Stage 4 [mg] 25.18 115.92 29.08 7.30 Stage 5 [mg]24.18 92.74 20.25 4.36 Stage 6 [mg] 6.82 16.64 5.33 1.27 Stage 7 [mg]1.17 4.76 1.32 0.31 MOC^(a)) [mg] 0.18 0.23 0.04 0.01 Sum [mg] 64.64259.29 65.69 15.66 Delivered mass 0.87 3.96 4.04 3.92 [g] FPD^(b), f))≤5 μm 55.1 218.8 52.8 12.4 [mg] FPD/Delivered 63.3 55.2 13.1 3.2 mass[mg/g] FPF^(c), f)) ≤5 μm 85.2 84.4 80.4 79.3 [%] GSD^(d), f)) 1.5 1.41.5 1.4 MMAD^(e), f)) [μm] 3.3 3.5 3.6 3.7 ^(a))MOC: Micro-OrificeCollector ^(b))FPD: Fine Partice Dose ^(c))FPF: Fine Partice Fraction;FPF is the FPD expressed as a percentage of the delivered dose ^(d))GSD:Geometric Standard Deviation ^(e))MMAD: Mass Median AerodynamicDistribution ^(f))After determination of the amount of drug (activecompound) deposited on the various stages FPD; FPF, GSD and MMAD werecalculated using the CITAS program, version 3.10.

From the results above, no significant changes were seen in theperformance of the solutions of different strength within the presentedrange, except the effect of the reduced concentration on the drugtotals.

GLP-Compliant 28-Day Inhalation Toxicity Study in Rats

In a GLP-compliant 28-day inhalation toxicity study in rats, conductedby Charles River Laboratories Preclinical Services, Tranent, Edinburgh,UK, the formulation described in Example 2 and dilutions thereof as wellas the vehicle were administered using the Pari eFlow® XL 30 nebulizerdevice (Pari Pharma, Starnberg, Germany) for 100 min per day for 28days, followed by a 2-week recovery period. Rats were treated withvehicle (0.6% [w/w] aqueous sodium chloride solution adjusted to pH 5.5with 1 M HCl pharmaceutical grade) or aerosols containing 0.15, 0.73 and1.63 mg/L ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-),corresponding to overall mean group achieved drug doses of 0, 11, 53 and119 mg/kg/day, respectively. Ten animals were used for the main toxicitystudy, and 5 additional animals for the recovery phase. Parametersassessed included clinical signs, body weights, food consumption,ophthalmic examination, clinical pathology, gross necropsy findings,organ weights and histopathologic examinations. Slightly reduced bodyweight gain was observed in males at 119 mg/kg/day compared to thevehicle control group. This was associated with reduction in foodconsumption in treated animals. There was a good recovery in body weightgain during the recovery period between day 28 and 42. There were noclinical signs or ophthalmic findings.

There were no findings in clinical pathology (haematology, coagulation,clinical chemistry and urinalysis investigations) which were consideredtoxicologically relevant. There were no treatment-related changes inorgan weights or gross findings following treatment with the drug.Histopathology of the larynx revealed focal minimal squamous metaplasiawithout cellular atypia of dysplasia in all groups. These changesresolved during the recovery period and were considered a non-adverse,adaptive response to mild irritation. Histopathology of the lungsrevealed minimal to moderate multifocal alveolar macrophage accumulationin all groups. This slight increase in macrophage accumulation wasconsidered a nonspecific response to the inhalation of highconcentrations of material exceeding the clearance capacity of the lung.In conclusion, the No Observed Adverse Effect Level (NOAEL) was 119mg/kg/day since the changes in body weight gain and food intake and thehistopathologic findings in the larynx were considered non-adverse.

GLP-Compliant 28-Day Inhalation Toxicity Study in Monkeys

In a GLP-compliant 28-day inhalation toxicity study in cynomolgusmonkeys, conducted by Charles River Laboratories Preclinical Services,Tranent, Edinburgh, UK, the formulation described in Example 2 anddilutions thereof as well as the vehicle were administered using thePari eFlow® XL 30 nebulizer device (Pari Pharma, Starnberg, Germany)with an oro-nasal inhalation mask for 60 min per day for 28 days,followed by a 2-week recovery period. Monkeys were treated with vehicle(0.6% [w/w] aqueous sodium chloride solution adjusted to pH 5.5 with 1 MHCl pharmaceutical grade) or aerosolized acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)corresponding to estimated mean achieved drug doses of 0, 11.2, 30.1 and112 mg/kg/day, respectively. In total five males and five females wereused for each dose of the toxicity study including recovery. Parametersassessed included clinical signs, body weights, electrocardiology,ophthalmic examination, clinical pathology, toxicokinetic parameters inthe plasma, gross necropsy findings, organ weights and histopathologicexaminations. There were no body weight changes, clinical signs,ophthalmic or electrocardiographic findings or changes in urinaryparameters attributable to drug treatment. In haematology and clinicalpathology, some minor, toxicologically insignificant changes wereobserved. None of these findings was considered toxicologicallyimportant, as the magnitude of responses was small, and most findingswere confined to one sex and/or showed evidence of regression when bothsexes were affected. Gross findings included increased lung weight inmales at 112 mg/kg/day, and one animal with a mottled appearance of thelungs and enlarged tracheobronchial lymph node. Histopathologic findingswere observed at all drug dose levels, including increased numbers ofalveolar macrophages, perivascular/peribronchiolar infiltrates, andgranular eosinophilic deposits in the lung and lymphoid hyperplasia inthe tracheobronchial lymph nodes, which showed complete recovery. Thesefindings demonstrated full reversibility during recovery, and wereconsidered to be due to inhalation of material exceeding normal lungclearance capacity, particularly at 112 mg/kg/day. In conclusion, NOAELin this study was 112 mg/kg/day, since the changes discussed above wereall considered non-adverse.

First-in-Man Study in Healthy Subjects to Investigate Safety andTolerability of Orally Inhaled Single Doses of a Formulation of AcetateSalt ofCyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)

In this randomised, double-blind, placebo-controlled, parallel-group(per dose level) dose-escalation study of inhaled single doses in 48healthy subjects in six dose groups of eight subjects each, conducted byInamed GmbH, Gauting, Germany, the controlled oral inhalation of theformulation described in Example 1 and dilutions thereof as well as theplacebo (0.5% [w/w] aqueous sodium chloride solution adjusted to pH 5.5with 1 M HCl pharmaceutical grade) occurred via the Pari eFlow® XL 30nebulizer device (Pari Pharma GmbH, Starnberg, Germany). The dose levelsand corresponding concentrations ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)(active compound) are summarized in Table 10.

TABLE 10 Preparation and Dose Volume of nebulizer solution ConcentrationDose Level 1 20 mg of drug in 4 mL solution  5 mg/mL Dose Level 2 60 mgof drug in 4 mL solution 15 mg/mL Dose Level 3 120 mg of drug in 4 mLsolution 30 mg/mL Dose Level 4 240 mg of drug in 4 mL solution 60 mg/mLDose Level 5 480 mg of drug in 6 mL solution 80 mg/mL Dose Level 6 960mg of drug in 12 mL 80 mg/mL solution

The volume of placebo nebulizer solution corresponded to that of theactive compound at a particular dose level. The duration of inhalationdepended on the total volume of nebulizer solution and ranged betweenseveral minutes and roughly one hour. Adverse events were tabulated andsummarized according to the current version of Medical Dictionary forRegulatory Activities.

Safety and Tolerability Results

No death, no serious adverse events (AE) and no other significant AEoccurred during the study. In total, 27 AEs, thereof 24treatment-emergent adverse events (TEAEs) were recorded in 13 subjects(27.1%). All of these subjects were on active compound. No AE wasreported for subjects having inhaled placebo solution. Regarding thenumber of AEs as well as their intensity and causal relationship to thestudy medication, a higher total number of AEs and related AEs werereported from the group inhaling the highest dose of active compound(960 mg; dose group 6) when compared to the other dose levels. Themajority of AEs and symptoms reported were related to the respiratorysystem, such as ‘cough’, ‘respiratory tract irritation’, increased mucusproduction or a transient decline in forced expiratory volume in thefirst second (FEV₁). The higher occurrence of respiratory symptoms andrespiratory AEs, especially in the groups inhaling higher doses ofactive compound, may possibly have been related to the long duration ofinhalation. These AEs may not necessarily have been related to theformulation of Example 1 and dilutions thereof itself, but may possiblybe procedural AEs. Regarding the local tolerability, there were threeinhalation-related adverse events (‘cough’). The overall tolerabilitywas judged as ‘very good’ or ‘good’ by the majority of subjects (97.9%).Throughout the study, the majority of the clinical laboratory valuesremained within the respective reference ranges. Most of the individualresults of physical examination, vital-signs measurements,electrocardiogram (ECG) recordings and lung-function results were withinthe commonly accepted clinical reference ranges. No time-dependentinfluence of active compound on safety parameters measured becameobvious. There was no relevant difference between the different dosegroups.

The above presented results of the First-in-Man study show that theformulation of Example 1 and dilutions thereof are highly suitable foraerosolization in a wide range of concentrations and applicable forinhalation administration in humans even at high concentrations (80mg/mL).

Effect of Inhalation Administration of Acetate Salt ofCyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) in anIn Vivo Model of Neutrophil Activation in the Rat

The purpose of this study (conducted by Envigo CRS Limited, Alconbury,Huntingdon, United Kingdom) was to evaluate the effects of acetate saltof cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation in a LPS/fMLP model of neutrophil activationin the rat.

Preparation of Formulations

The formulation for test animal group 1 (vehicle) was 0.5% (w/v) salineadjusted to a pH of 5.5 with 1 M HCl and filtered through a 0.2 μmfilter.

The formulations for test animal groups 2-4 were prepared as follows:

The appropriate amount of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) wasweighed and the appropriate amount of 0.5% (w/v) saline, pH 5.5, wasadded to produce a formulated concentration of 83.28 mg/mL.

The pH of the final solution was adjusted to 5.5 with 1 M NaOH.

An appropriate amount of 83.28 mg/mL of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) wasadded to an appropriate amount of vehicle to produce a formulation of15.62 mg/mL.

Finally, an appropriate amount of 15.62 mg/mL of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) wasadded to an appropriate amount of vehicle to produce a formulation of5.21 mg/mL. The three formulation solutions (83.28 mg/mL, 15.62 mg/mL,5.21 mg/mL) were then filtered through a 0.2 μm filter.

The formulations were prepared 1 day prior to dosing and stored at 2-8°C. in the dark until the day of use when they were removed from thefridge and maintained at room temperature 25° C.) and gently shaken forat least 1 h prior to dosing.

Table 11 summarizes the concentrations of the prepared formulations fortest animal groups 1-4 taking into account a ratio of acetate salt/freebase ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) being1.041. Thus, these formulations correspond to the formulation describedin Example 1 and dilutions thereof.

TABLE 11 Nominal Formulated Group concentration of drug concentration ofdrug 1 (control)  0 mg/mL    0 mg/mL 2  5 mg/mL  5.21 mg/mL 3 15 mg/mL15.62 mg/mL 4 80 mg/mL 83.28 mg/mL

Procedure for Inhalation Treatment Cohort

Animals were challenged with aerosolized LPS (lipopolysaccharide, 1mg/mL) for 30 min. Approximately 3 h following end of LPS challenge,animals were administered either vehicle or formulations as describedabove of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) byinhalation (Groups 1-4) over a 30 min period using an eFlow® 30 XLelectronic nebulizer (Pari Pharma GmbH, Starnberg, Germany; snout onlyexposure). All animals were dosed with fMLP (N-formyl-Met-Leu-Phe, 5mg/kg) approximately 4 h post completion of the LPS challenge via theintratracheal route under transient gaseous anaesthesia at a dose volumeof 1 mL/kg. Approximately 2 h after fMLP administration, animals wereterminated and a bronchoalveolar lavage (BAL) was performed to evaluateinflammatory cell infiltrate and neutrophil elastase activity. Theprocedure for the inhalation treatment cohort is summarized in Table 12.

TABLE 12 Inhalation treatment; nominal Target Treatment Lung doseNeutrophil Whole body concentration exposure dose (mg/kg) elastaseAnimal challenge of drug level (mg/kg) (assuming 10% activation numbersGroup (30 min) (mg/mL) (μg/L)* free base) deposition) (1 mL/kg, i.t.)(PD) 1 LPS  0 mg/mL — — — fMLP 10 (1 mg/mL) (vehicle) (5 mg/kg) 2 LPS  5mg/mL 13.4 0.3 0.03 fMLP 10 (1 mg/mL) (5 mg/kg) 3 LPS 15 mg/mL 134 3 0.3fMLP 10 (1 mg/mL) (5 mg/kg) 4 LPS 80 mg/mL 1340 30 3 fMLP 10 (1 mg/mL)(5 mg/kg) *Exposure level (μg/L) based on a 250 g rat. LPS in 0.9% (w/v)saline fMLP in 1% DMSO in saline Male rat, Crl:CD (SD), Charles RiverLaboratories, Wilmington, Massachusetts, USA All animals were terminatedfor bronchoalveolar lavage approximately 6 h after LPS exposure.

${{Delivered}\mspace{14mu}{dose}\mspace{14mu}\left( {{µg}\text{/}{kg}} \right)} = \frac{{C\left( {{µg}\text{/}L} \right)} \times {RMV}\mspace{14mu}\left( {L\text{/}\min} \right) \times D\mspace{11mu}\left( \min \right)}{{BW}\mspace{14mu}({kg})}$where C = Concentration  in  air  inhaledRMV = Respiratory  minute  volume, calculated  from  the       formula:D = Duration  of  exposure  in  min  BW = BodyweightRef.  1:  D.  J.  Alexander, C.  J. Collins, D.  W.  Coombs  et  al., Association  of  Inhalation  Toxicologists  (AIT)  working  partyrecommendation  for  standard  delivered  dose  calculation  andexpression  in  non-clinical  aerosol  inhalation  toxicology  studieswith  pharmaceuticals; Inhal.  Tox., 2008, 20, 1179-1189.

The doses of fMLP and the time points were chosen based upon publisheddata (see e.g. S. Yasui, A. Nagai, K. Aoshiba et al., Eur. Respir. J.,1995, 8, 1293; T. Yang, J. Zhang, K. Sun et al., Inflamm. Res. 2012, 61,563; R. Corteling, D. Wyss, A. Trifilieff, BMC Pharmacology, 2002, 2, 1)and experience at Envigo CRS Limited. The dose selection of the drug wasbased on a previous intratracheal study at Envigo CRS Limited. In thisprevious study, intratracheal doses of 0.03 to 3 mg/kg of the drug werefound to be efficacious in this animal model.

No test item related clinical signs were observed between the dosingperiod and study termination. Two animals of group 1 (vehicle) died justafter fMLP administration due to deep anaesthesia.

Bronchoalveolar Lavage (BAL)

Following confirmation of death, the trachea of an animal was isolated,the tracheal cannula inserted and secured in place, and the airway waslavaged with 3 mL of phosphate buffered saline (PBS). The lavage wasrepeated twice and in total, three lots of 3 mL of PBS were used. Thefirst lavage aliquot containing cells was placed into a 15 mL centrifugetube on wet ice (Tube A). The BAL fluid pooled from the second twolavages was placed into a second tube (Tube B). Tube A and B were placedon wet ice until centrifuged. Centrifugation was performed at 800×g for10 min at ca. 4° C. and the supernantant was harvested. Until neutrophilelastase analysis supernantant was stored at ca. 80° C.

Neutrophil Elastase Activity

The two aliquots of BAL supernatant were analyzed for neutrophilelastase activity as follows: 120 μl of BAL supernatant from aliquot 1was transferred to a 96 well plate (Corning #3650). In parallel, adilution range from 1.6 to 0.025 mU/well of a commercial humanneutrophil elastase (hNE, Serva #20927.01) was prepared. 120 μl of eachhNE dilution were transferred in duplicate to the 96-well plate. Tostart the enzymatic reaction, 80 μl of a fluorescent peptide substrate(MeOSuc-Ala-Ala-Pro-Val-AMC) at a final concentration of 500 μM wasadded to each well and the plate was immediately placed in the victor2vfluorescent reader pre-warmed at 37° C. Fluorescence (λexc. 485 nm, λem.535 nm) was recorded for 2 h at 37° C. Enzyme initial velocity (RFU/min)of all samples was calculated and converted in mU/mL hNE equivalent,using the linear regression equation obtained from the plot (RFU/min vsmU/mL of hNE dilution) of the human neutrophil elastase standard range.The assay was repeated using BAL supernatant from aliquot 2. Theneutrophil elastase data reported is a mean of the neutrophil elastaseactivity from both aliquots. The neutrophil elastase activity in the BALfluid is presented in Table 13 and FIGS. 1 and 2 .

TABLE 13 Group Inhalation administation NE activity (mU/mL eq.) 1vehicle/fMLP 0.25 ± 0.02   2 drug (0.3 mg/kg)/fMLP 0.10 ± 0.01**** 3drug (3 mg/kg)/fMLP 0.11 ± 0.01**** 4 drug (30 mg/kg)/fMLP 0.07 ±0.01**** Values rounded, precision may not be as displayed. Data isexpressed as mean ± s.e.m. ****p < 0.0001 when compared to the vehicle(inhalation)/fMLP treated group.

The drugcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered at doses of 0.3, 3 and 30 mg/kg by inhalation route 3 hpost LPS challenge and 1 h prior to fMLP challenge significantlyinhibited neutrophil elastase activity in BAL fluid harvested 6 h postLPS challenge.

Phase-1b Study to Investigate Safety, Tolerability and Pharmacokineticsof Orally Inhaled Single Doses of Acetate Salt ofCyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-) inPatients with Cystic Fibrosis

In this randomised, double-blind, placebo-controlled, parallel-group(per dose level) dose-escalation study, the safety and tolerability ofsingle ascending doses (SAD) of acetate salt ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by inhalation in patients with cystic fibrosis (CF) wasinvestigated. Additionally, the pharmacokinetics of the drug following asingle ascending dose in plasma and sputum as well as itspharmacodynamic effect on neutrophil elastase activity in sputum wereevaluated.

Treatment

In this study, conducted by Inamed GmbH, Gauting, Germany, 24 subjectswith cystic fibrosis, who fulfilled all the inclusion criteria and inwhom no exclusion criterion was present, were included and receivedrandomised treatment. They were grouped into 3 dose groups of 8 subjectseach. 6 subjects received the formulation described in Example 1 anddilutions thereof and 2 subjects received placebo (0.5% [w/w] aqueoussodium chloride solution adjusted to pH 5.5 with 1 M HCl pharmaceuticalgrade). The oral inhalation of the above formulation as well as theplacebo occurred via the Pari eFlow® XL 30 nebulizer device (Pari PharmaGmbH, Starnberg, Germany). The dose levels and correspondingconcentrations ofcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)(active compound) are summarized in Table 14.

TABLE 14 Preparation and Dose Volume of nebulizer solution ConcentrationDose Level 1 80 mg of drug in 4 mL solution 20 mg/mL Dose Level 2 160 mgof drug in 4 mL solution 40 mg/mL Dose Level 3 320 mg of drug in 4 mLsolution 80 mg/mL

The volume of placebo nebulizer solution corresponded to that of theactive compound at a particular dose level. The duration of inhalationwas estimated to be between 7 and 20 min. Adverse events were tabulatedand summarized according to the current version of Medical Dictionaryfor Regulatory Activities.

Safety and Tolerability Results

No death, no serious adverse events (AE) and no other significant AEoccurred during the study. There were no inhalation-related local AEs.None of the 24 subjects terminated the study early because of AE.

In total, 6 AEs, all of them treatment-emergent adverse events (TEAEs),were recorded in 6 subjects. The most frequently reported terms were‘dizziness’ and ‘headache’ with 2 events each. All 6 TEAEs were regardedas being not related to the active compound. None of the TEAEs was ratedas severe. AE duration was transient. All 6 AEs resolved withoutsequels. Regarding the number of AEs as well as their intensity andcausal relationship to the study medication, no obvious differencebetween the dose levels became apparent. Throughout the study, themajority of clinical laboratory values as well as ECG and vital signsresults remained within the respective reference ranges. Any deviatingfindings were clinically not relevant and well in line with the extentof deviations usually observed in studies with CF patients. The majorityof results from lung function tests were as expected of patients withCF.

Pharmacokinetic Assessments

The following pharmacokinetic parameters were assessed forcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)(active compound):

Plasma:

AUC_(0-∞), AUC_(0-∞)/D, C_(max), C_(max)/D of active compound in plasmaas primary variables and t_(max), t_(1/2), λ_(z), AUC_(0-tlast) ofactive compound in plasma as secondary variables.

Sputum:

Concentrations of active compound in sputum were calculated and C. andAUC_(0-tlast) were determined.

Sampling and Sample Processing for Plasma Samples

Blood samples for the determination of the plasma concentrations ofactive compound were taken at the following points in time:

On Day 1 at pre-dose, 10 min, and 0.5 h, 1 h, 2 h, 3 h, 4 h, 6 h, 8 h,12 h, 15 h, and on Day 2 at 24 h post dose (after start of inhalation).

The samples were collected in closed K3-EDTA plasma sampling tubes(Monovette® Sarstedt, Germany). The accepted time-interval for samplehandling procedures for each individual study sample, i.e., the timebetween sample collection and sample centrifugation had not to exceed 60min, and the time between the end of centrifugation and sample freezinghad also not to exceed 60 min.

The samples were centrifuged at approximately 4° C. (±2° C.) at 2200×gfor 15 min. The resulting plasma supernatant was then transferred into 2polypropylene tubes (1^(st) aliquot of at least 1 mL and backup) andfrozen in upright position below 20° C. Samples were stored in a freezerunder continuous temperature control below 20° C. from the day aftersample collection until shipment (dry ice with a thermo-logging device)to the bioanalytical site (Pharmacelsus GmbH, Saarbrucken, Germany).

Sampling and Sample Processing for Sputum Samples

Spontaneous sputum samples for PK assessments were collected during thefollowing time intervals/periods:

On Day −1 as soon as possible after the subject's arrival at the studysite (blank PK), on Day 1 in the period of time between 1 h and 3 hafter start of inhalation, and in the morning of Day 2, at approx. 24 hafter start of inhalation. Moreover, any spontaneous sputumexpectoration between 0-1 h after start of inhalation was collected.

Spontaneous sputum samples were collected in polystyrene Petri dishesand put on ice immediately. All processing steps had to be performedwith cold reagents and on ice whenever possible. Plugs were separatedfrom saliva, but the latter was not discarded. If a sputum sample wastaken for both PK and neutrophil elastase activity in sputum analysis,the plug was split in 2 approximately equal parts. One part wasprocessed for PK analysis the other one for evaluation of neutrophilelastase activity in sputum.

For PK processing, the resulting plug was split in approximately 2 equalparts. These were transferred into 2 transfer tubes for PK evaluation(SP1) and PK Backup (SP3). Up to 1.0 mL saliva was transferred intoanother transfer tube for PK evaluation (SP2). The remaining saliva inthe Petri dish was discarded. The weight of the sputum samples (SP1+SP3)was determined and all samples were immediately put on dry ice forfreezing. After freezing, samples were stored in an upright position at−80±10° C.

The sputum and saliva samples for PK evaluation (SP1+SP2) were shippedto Pharmacelsus GmbH on dry ice. The backup sputum samples (SP3) werestored at −80±10° C. at Inamed GmbH, Gauting, Germany.

Bioanalytical Methods

For the analysis of active compound in plasma, a validated and highlysensitive liquid chromatography tandem mass spectrometry (LC-MS/MS)method was used.

For the analysis of active compound in sputum a validated or, where notavailable, ‘fit-for-purpose’ highly sensitive liquid chromatographytandem mass spectrometry (LC-MS/MS) method was to be used.

The bioanalytical procedures were performed according to current GoodLaboratory Practice (GLP) regulations, US Food and Drug Administration(FDA) and EMA validation requirements for bioanalytical assays and wereoutlined in applicable SOPs including regulations for routine analysisand general regulations for analysis.

Derived Pharmacokinetic Parameters

The pharmacokinetic parameters mentioned above were calculated based onactual blood and sputum sampling times using non-compartmentalprocedures.

Plasma Concentration-Time Curves and Derived Pharmacokinetic Parametersof Cyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)

The mean plasma concentration-time curves of active compound are shownin FIG. 3 .

The curves of active compound show typical profiles and similar to theresults of healthy subjects inhaling comparable doses of activecompound, its first plasma concentrations were detected early on inpatients with CF.

Plasma concentrations of active compound increased to reach theirrespective peaks (t_(max), mean) at 1.3 h in dose group 1, at 1.5 h indose group 2 and at 2.3 h in dose group 3. Thereafter, plasmaconcentrations of active compound declined with a mean terminalhalf-life of 4.1 h in dose group 1, 3.5 h in dose group 2 and 3.8 h indose group 3. 24 hours after inhalation, active compound could still bedetected in subjects of all dose groups, with mean plasma concentrationsof 2.8 ng/mL in dose group 1, 4.4 ng/mL in dose group 2 and 15.2 ng/mLin dose group 3.

Sputum Concentration ofCyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)

The mean sputum concentrations of active compound are shown in FIG. 4 .

Already at the first sputum sampling time point after inhalation (0-1 hsampling interval), active compound was detected in the majority ofsubjects across all dose groups. The concentration of active compound insputum increased by dose, with the highest mean (SD) concentrationvalues of 0.6 (0.79) g/L (dose group 1, 1-3 h sampling interval), 1.1(1.23) g/L (dose group 2, 0-1 h sampling interval) and 1.8 (1.81) g/L(dose group 3, 0-1 h sampling interval). Active compound could be stilldetected in sputum across subjects of all dose groups at 24 h afterinhalation.

Pharmacodynamic Assessments

The pharmacodynamic effect of ascending single doses of active compoundwas investigated by evaluation of neutrophil elastase (NE) activity insputum using FRET (fluorescence resonance energy transfer) assay afterPBS aliquotation and freezing.

Sampling and Sample Processing for Sputum Samples

Spontaneous sputum samples for evaluation of neutrophil elastase (NE)activity were collected during the following time intervals/periods:

On the day of screening, if possible, for baseline assessment of NEactivity, on Day −1 as soon as possible after the subject's arrival atthe study site for baseline assessment of NE activity, on Day 1 in theperiod of time between 1 h and 3 h after start of inhalation, and in themorning of Day 2, at approx. 24 h after start of inhalation.

Spontaneous sputum samples were collected in polystyrene Petri dishesand put on ice immediately. All processing steps had to be performedwith cold reagents and on ice whenever possible. Plugs were separatedfrom saliva, but the latter was not discarded. If a sputum sample wastaken for both PK and NE activity in sputum analysis, the plug was splitin 2 approximately equal parts. One part was processed for evaluation ofNE activity in sputum the other one for PK analysis, as described above.

For evaluation of NE activity in sputum, at first, the weight of thesputum sample was determined and 8 mL cold phosphate-buffered saline(PBS) per gram purified sputum was added. The sample was vortexed for 30s at room temperature (15-24° C.). Then, the soluble fraction wasseparated from the sputum pellet by centrifugation (10 min at 1000×g and4° C.). The supernatant was transferred to a fresh tube.

Remaining insoluble particles were separated by a second centrifugationstep at 3500×g for 15 min at 4° C. If the supernatant was not aliquotedimmediately, the PBS/sputum supernatant was transferred to a fresh tube.

10 aliquots of 50 μL PBS/sputum were transferred into transfer tubes forevaluation. Moreover, 4 aliquots of 50 μL PBS/sputum were transferredinto transfer tubes as backup. In case there was a leftover, theremaining volume was immediately transferred into a transfer tube forevaluation. Any leftover exceeding 2.0 mL was discarded.

Samples were frozen immediately and stored in an upright position at−80±10° C. PBS/Sputum samples were shipped to MLM Medical Labs GmbH,Mönchengladbach, Germany, on dry ice. PBS/Sputum backup samples werestored at −80±10° C. at Inamed GmbH, Gauting, Germany.

Bioanalytical Methods

The NE activity in sputum was evaluated by using a FRET elastase assayadapted from an assay described in Nature Protocols, 2008, 3, 991 (B.Korkmaz, S. Attucci, M. A. Juliano et al.) and validated by MLM MedicalLabs GmbH, Mönchengladbach, Germany.

The assay is based on the reaction of human neutrophil elastase with thesubstrate 2Abz-Ala-Pro-Glu-Glu-Ile-Met-Arg-Arg-Gln-Tyr(3NO₂)—OH(GeneCust Europe S.A., Ellange, Luxembourg, #P160301-SY452824). Byadding the substrate solution the enzyme reaction is started and theelastase present in the samples reacts with added substrate. The productof the reaction is detected through fluorescence measurement and theinitial reaction velocity is determined. The concentration of activeelastase in the unknown samples is back-calculated using the calibrationcurve calculated from the standards.

An elastase reference stock solution (human neutrophil elastase [ELA2],Höltzel Diagnostika, Köln, Germany, #PN31255) with known enzymaticactivity was prepared. The exact active elastase enzyme concentrationwas determined by titration against an alpha 1-antitrypsin solution(Athens Research and Technology, Athens, USA, #16-16-011609), with adefined concentration. The active elastase reference stock solution wasadjusted by addition of PBS to a concentration of 3000 nM and aliquotswere stored at −80° C.

The FRET assay was performed in 96 well white microtiter plates in totalvolume of 100 uL per well. PBS sputum dilutions of 1:5, 1:50 and 1:100in elastase reaction buffer (50 mM HEPES, pH 7.4, 750 mM NaCl, 0.05%[v/v] NP-40) were prepared, 5 uL loaded per well and 90 uL elastasereaction buffer were added. A 5 mM substrate stock solution of2Abz-Ala-Pro-Glu-Glu-Ile-Met-Arg-Arg-Gln-Tyr(3NO₂)—OH in 30% (v/v)N,N-dimethylformamide/water was prepared and further diluted to 400 nMby addition of elastase reaction buffer. 5 uL of the 400 nM substratesolution were added to the wells and the reaction velocity V_(i) wasdetermined by FRET at Exc:320 nm-Emiss:420 nm. The concentration ofactive elastase in PBS sputum was determined by comparison of reactionvelocities of defined dilutions from the active elastase reference stocksolution.

The analytical measuring range of this method was 115.00-2880.00 ng/mL.Intra- and inter-assay coefficients of variation were ≤17.14% and≤8.96%, respectively.

Active Neutrophil Elastase in Sputum

Mean concentrations of active NE are shown in FIG. 5 .

Active NE concentration results were highly variable throughout. Whenmeasured by FRET assay, mean (SD) concentrations of active NE in sputumon Day −1 (predose) were 18823.6 (19758.72) ng/mL for dose level 1,9548.7 (6222.55) ng/mL for dose level 2 and 10480.7 (10528.00) ng/mL fordose level 3. The mean (SD) active NE concentration for placebo subjectsat pre-dose was 46711.8 (48456.86) ng/mL.

The concentration of active NE in sputum strongly decreased in subjectsafter inhaling active compound. Mean (SD) concentration values at 1-3 hafter inhalation were 612.5 (1218.62) ng/mL for dose level 1 and 115.0(0.00) ng/mL for dose levels 2 and 3. The mean (SD) value for subjectsinhaling placebo at 1-3 h after inhalation was 34370.0 (19988.11) ng/mL.

Mean (SD) active NE concentrations in sputum at 24 h after inhalationwere 1467.7 (2154.43) ng/mL for dose level 1, 16849.4 (23518.56) ng/mLfor dose level 2 and 7712.5 (11394.04) ng/mL for dose level 3. The mean(SD) active NE concentration at 24 h for placebo subjects was 19338.8(9062.32) ng/mL.

Pharmacodynamic Conclusions

The mean concentration of active NE in the sputum of patients with CFstrongly decreased after single-dose inhalation of active compoundadministered by a formulation described in Example 1. In the dose rangeexamined, the extent of this response appeared to be independent of thedose administered. Although difficult to judge due to high datavariability, the mean concentration of active NE in sputum apparentlyreturned to baseline levels at 24 h after inhalation of active compound.The inhalation of placebo solution had no distinct effect on the meanconcentration of active NE in sputum. These clinical results show thatcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-)administered by a formulation described in Example 1 inhibits NE in thesputum of patients with CF.

The invention claimed is:
 1. A method of treating diseases or conditionsof the lungs being mediated by or resulting from human neutrophilelastase activity, the method comprising administering an effectiveamount in a subject in need thereof of a pharmaceutical compositioncomprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; wherein OctG is(S)-2-aminodecanoic acid; ^(D)Pro is D-proline; wherein thepharmaceutical composition further comprises sodium chloride; andoptionally one or more pharmaceutically acceptable diluents, excipientsor carriers; and wherein the pharmaceutical composition is administeredto the subject as pharmaceutical aerosol for pulmonary administrationcomprising a dispersed liquid phase and a continuous gas phase, whereinthe dispersed liquid phase (a) comprises aqueous droplets comprising theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; (b) has a mass mediandiameter from about 1.5 um to about 5 um; and (c) has a droplet sizedistribution having a geometrical standard deviation from about 1.2 toabout 1.7; and wherein said administration inhibits neutrophil elastaseactivity in said subject.
 2. The method according to claim 1, whereinthe diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity are pulmonary diseases.
 3. Themethod according to claim 2, wherein the diseases or conditions of thelungs being mediated by or resulting from human neutrophil elastaseactivity are pulmonary diseases consisting of alpha-1 antitrypsindeficiency (AATD), cystic fibrosis (CF), non-cystic fibrosisbronchiactasis (NCFB), or chronic obstructive pulmonary disease (COPD),or infections of the lungs causing diseases or conditions of the lungs,being mediated by human neutrophil elastase activity.
 4. The methodaccording to claim 3, wherein the pulmonary disease is non-cysticfibrosis bronchiactasis (NCFB) or cystic fibrosis (CF).
 5. The methodaccording to claim 3, wherein the pulmonary disease is cystic fibrosis(CF).
 6. The method according to claim 1, wherein the active compound iscombined with the counter ion acetate.
 7. The method according to claim1, wherein the aerosol being emitted from an aerosol generator at a rateof at least about 0.1 mL dispersed liquid phase per minute.
 8. Themethod according to claim 1, wherein the aerosol being emitted from anaerosol generator at a mean delivery rate of at least about 0.8 mg ofthe active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; per minute.
 9. The methodaccording to claim 1, wherein the pharmaceutical composition is a liquidpharmaceutical composition for preparing the aerosol comprising theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; in a concentration withina range from about 4 mg/mL to about 100 mg/mL.
 10. The method accordingto claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 0.1 and about 10000 mg/day.
 11. Themethod according to claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 0.001 and about 100 mg/kg.
 12. Themethod according to claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 5 and about 1000 mg/day.
 13. The methodaccording to claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose of about 20, about 60, about 120, about 240, about 480or about 960 mg/day.
 14. The method according to claim 1, wherein theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 80 and about 320 mg/day.
 15. The methodaccording to claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose of about 80, about 160, or about 320 mg/day.
 16. Themethod according to claim 1, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof is administered to thesubject by oral inhalation.
 17. A method of treating diseases orconditions of the lungs being mediated by or resulting from humanneutrophil elastase activity, the method comprising administering aneffective amount in a subject in need thereof of a pharmaceuticalaerosol for pulmonary administration comprising a dispersed liquid phaseand a continuous gas phase, wherein the dispersed liquid phase (a)comprises aqueous droplets comprising the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; wherein OctG is(S)-2-aminodecanoic acid; ^(D)Pro is D-proline; (b) has a mass mediandiameter from about 1.5 μm to about 5 μm; and (c) has a droplet sizedistribution having a geometrical standard deviation from about 1.2 toabout 1.7; and wherein said administration inhibits neutrophil elastaseactivity in said subject.
 18. The method according to claim 17, whereinthe diseases or conditions of the lungs being mediated by or resultingfrom human neutrophil elastase activity are pulmonary diseasesconsisting of alpha-1 antitrypsin deficiency (AATD), cystic fibrosis(CF), non-cystic fibrosis bronchiactasis (NCFB), or chronic obstructivepulmonary disease (COPD), or infections of the lungs causing diseases orconditions of the lungs, being mediated by human neutrophil elastaseactivity.
 19. The method according to claim 18, wherein the pulmonarydisease is non-cystic fibrosis bronchiactasis (NCFB) or cystic fibrosis(CF).
 20. The method according to claim 18, wherein the pulmonarydisease is cystic fibrosis (CF).
 21. The method according to claim 17,wherein the active compound is combined with the counter ion acetate.22. The method according to claim 17, wherein the aerosol being emittedfrom an aerosol generator at a rate of at least about 0.1 mL dispersedliquid phase per minute.
 23. The method according to claim 17, whereinthe aerosol being emitted from an aerosol generator at a mean deliveryrate of at least about 0.8 mg of the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; per minute.
 24. The methodaccording to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 0.1 and about 10000 mg/day.
 25. Themethod according to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 0.001 and about 100 mg/kg.
 26. Themethod according to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 5 and about 1000 mg/day.
 27. The methodaccording to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose of about 20, about 60, about 120, about 240, about 480or about 960 mg/day.
 28. The method according to claim 17, wherein theactive compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose between about 80 and about 320 mg/day.
 29. The methodaccording to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject at a dose of about 80, about 160, or about 320 mg/day.
 30. Themethod according to claim 17, wherein the active compoundcyclo(-OctG-Glu-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Lys-Tyr-^(D)Pro-Pro-); orany pharmaceutically acceptable salt thereof; is administered to thesubject by oral inhalation.
 31. The method according to claim 17,wherein the pharmaceutical aerosol further comprises one or more osmoticagents.
 32. The method according to claim 31, wherein the one or moreosmotic agents comprise sodium chloride.